bluuebunny/crossref_metadata_2025_split huggingface dataset:
Genes Dysregulated in a Murine Model of Leukemogenesis Comprise a Signature for Identification of Therapeutics in Humans.
Authors: Hassane Duane C , Ashton John M , Guzman Monica L , Jordan Craig T. 
Published: October 2019

Abstract
The identification of genes and pathways that are critical for the development and maintenance of leukemia is important for designing new anti-leukemia therapeutics. However, selection of aberrantly regulated genes and pathways and the translation of these features into targeted anti-cancer drugs represent a significant challenge. Studies have characterized the various oncogenic lesions associated with leukemia, and have shown that specific combinations of mutations are sufficient to induce malignancy in model systems. Thus, we sought to identify the gene signature arising from the introduction of cooperating oncogenic mutations into non-malignant cells. We hypothesized that compounds capable of maximally antagonizing the aberrant expression of this subset of genes would represent potential anti-leukemia therapeutics. To test this hypothesis, we turned to the use of murine genetics and chemical genomics. To this end, we adapted mouse systems created by retroviral transduction of primary marrow cells with the BCR-ABL translocation alone or in combination with the NUP98-HOXA9 translocation. This approach creates authentic models of chronic and blast crisis myeloid leukemia (CML and bcCML, respectively), which has been shown to closely resemble human disease. Next, we employed flow cytometry to separate the transduced marrow into purified primitive lin- populations of either normal, BCR-ABL+, or BCR-ABL+/NUP98-HOXA9+ cells. Importantly, lin- cells were used to focus specifically on those genes most relevant to leukemic stem and progenitor populations. Microarray gene expression profiling was performed on these purified populations in six independent experiments using the Affymetrix Mouse 430 2.0 platform. Gene expression signatures for each transduced population relative to normal lin- cells were obtained. Next, in silico screening for agents that antagonize these signatures was performed comparing the resulting signatures to a local microarray compendium that included the CMap dataset. Gene expression signatures arising from the introduction of BCR-ABL alone identified traditional chemotherapeutics including cytarabine, etoposide, idarubicin – agents considered less effective against leukemic stem cell populations – as well as agents acting along the PI3 kinase pathway. Surprisingly, drugs shown to antagonize the gene signature derived by the introduction the two oncogenic mutations (BCR-ABL and NUP98-HOXA9) included 15-delta-prostaglandin-J2, celastrol, MG- 132, and parthenolide. Each of these drugs eradicate malignant stem and progenitor cells, while sparing normal cells. Thus, the murine-derived signature successfully identified drugs highly relevant to human disease. Our studies therefore demonstrate the utility of combining gene expression-based drug screening and mouse cancer genetics to reveal novel anti-leukemia compounds in humans. DCH and JMA contributed equally.
Lmo2 Induces T-Cell Leukemia with Epigenetic Deregulation of CD4.
Authors: Cleveland Susan , Dave Utpal P. 
Published: October 2019

Abstract
The LIM domain Only-2 (Lmo2) oncogene is deregulated in the majority of human T-cell acute lymphoblastic leukemias (T-ALL). Enforced, constitutive expression of Lmo2 in transgenic mice using the T-cell-specific Cd2 promoter/enhancer induces T-ALL with median latency of 200 days and 100% penetrance. These transgenic mice have a pre-leukemic differentiation block at the double negative stage (CD4− CD8−) of T-cell differentiation. Despite this, T-ALLs in these mice arise from various stages of T-cell differentiation. To explore the relationship between T-cell development and Lmo2 oncogene function, we established cell lines from four independent T-cell tumors representing the double negative (DN), double positive (DP), and single positive (SP) stages. Two of the lines showed variegated expression of CD4 antigen. One line, 32080, was striking in that CD4 was expressed in 50% of cells whereas CD8 was uniformly present. The cell line expressed markers consistent with the intermediate single positive stage (ISP) to double positive (DP) stage transition. We sorted the 32080 line for highly CD4 positive and negative cells and observed them in culture. After one week, both sorted populations showed variegated CD4 expression. We determined CD4 expression was regulated at the transcriptional level and not due to post-transcriptional effects and that cell replication was necessary to re-establish CD4 variegation. Importantly, variegated CD4 expression was dependent on Lmo2 because its knockdown resulted in decreased CD4 expression. HDAC inhibition by trichostatin A had the reciprocal effect of inducing more CD4 expression suggesting that chromatin modification was playing a role. The CD4 locus was analyzed by chromatin immunoprecipitation and histone H3 methylation patterns consistent with silencing were found in the CD4 negative sorted cells but not in the CD4 positive population. The 32080 cell line is a striking model of ISP to DP T-cell plasticity and invokes a novel mechanism by which Lmo2 functions.
Development of a Preclinical Model for An Erythropoietin Deficient Anemia
Authors: Zeigler Brandon , Qin Wenning , Vajdos Janis , Loverro Linda , Niss Knut 
Published: October 2019

Abstract
The production of red blood cells is tightly controlled by the growth factor erythropoietin (EPO). Mice lacking a functional EPO gene die in-utero due to the complete absence of definitive red blood cells. Thus, studying the role of EPO in adult animals has been limited to the use of artificial anemia models such as phenylhydrazine (PHZ) treatment or phlebotomy. In addition, injections of rHuEPO have been shown to be a valuable tool to study erythropoiesis. However, these approaches have limited use as preclinical models since they do not recapitulate an anemia due to EPO deficiency as it is seen in patients with chronic kidney disease (CKD). These patients develop a chronic anemia due to the lack of a sufficient EPO gene expression response. In order to obtain a preclinical model for an EPO deficient anemia we developed a mouse model in which the EPO gene is flanked by two loxP sites (flox allele). In addition, these animals carry the Cre-ERT2 transgene which allows for a tamoxifen controlled activation of the Cre recombinase. Thus, these mice enable the inactivation of the EPO gene in adult animals by tamoxifen administration.
After induction of Cre activity, mice heterozygous for the EPO allele (EPOko/flox) develop a moderate to severe anemia within 3 weeks. Although anemic, these mice display only low levels of EPO in their serum. However, within 3 months after induction of the knock-out the anemia resolves. This indicates that the Cre mediated excision of the EPO gene is insufficient and some EPO expressing cells remain. This mimics the clinical situation of patients with CKD.
In order to study the response of EPO deficient mice to a severe anemia we treated animals with phenylhydrazine and observed the recovery of these animals. Interestingly, EPO deficient animals, although slightly delayed, fully recover from the induced anemia. Although the measured sEPO levels are significantly lower compared to control animals an increase in splenic erythropoiesis was observed in these animals. This indicates that the hypoxic response triggered by a sudden severe anemia is activating pathways other than EPO that promote erythropoiesis. Taken together, these results show that this novel mouse model will be useful tool as a preclinical model for the anemia associated with CKD. In addition, this model will allow the study of stress erythropoiesis under EPO limiting conditions. The Pfizer Institutional Animal Care and Use Committee reviewed and approved the animal use in these studies. The animal care and use program is fully accredited by the Association for Assessment and Accreditation of Laboratory Animal Care, International
Chk1 Haploinsufficiency Results in Anemia and Defective Erythropoiesis
Authors: Boles Nathan C , Peddibhotla Sirisha , Chen Alice , Rosen Jeffrey , Goodell Margaret A. 
Published: October 2019

Abstract
Erythropoiesis is a highly regulated and well-characterized developmental process responsible for providing the oxygen transport system of the body. However, besides erythropoietin’s role, few of the mechanisms involved in this process have been elucidated. Checkpoint Kinase 1 (Chk1) is best known for its role in the cell cycle and DNA damage pathways. Chk1 expression in the hematopoietic system has been demonstrated to be restricted to T-cells, erythroid cells and the hematopoietic stem cell. Interestingly, a lack of Chk1 has recently been shown to disrupt T-cell differentiation (Zaugg et al. PNAS 2007) Here, we show that haploinsufficiency of Chk1 leads to a 30% increase in the incidence of anemia in mice. Furthermore, the anemic Chk1 +/− mice exhibit distorted spleen and bone marrow architecture, and an increase in erythroid progenitors (MEP, stages I, and II). In young non-anemic Chk1 +/− mice, a decrease in enucleated cells of the erythroid lineage is observed, suggesting a role for Chk1 in proper erythroid enucleation. In addition, a decrease in Chk1 RNA has been observed in patients with refractory anemia with excess blasts (Pellagatti et al. Blood, 2006), further supporting an important role for Chk1 status in clinical anemia. Finally, Chk1 has recently been show to affect the regulation of Fancd2, one of the gene’s responsible for Fanconi anemia. These results indicate Chk1 plays a part in maintaining the balance between erythroid progenitors and the enucleated erythroid cells during differentiation.
The Impact of Rhg-CSF on the LFA-1 Conformation of CD4+T Cells in Mobilized Hematopoietic Stem Cell Allografts
Authors: Gao Chunji , Chen Weihua , Wang Fei , Li Meng , Zhu Haiyan , Da Wanming 
Published: October 2019

Abstract
In hemotopoietic stem cell transplantation (HSCT), the primary effects of G-CSF on cells of the hemotopoietic system include stimulation of proliferation and differentiation of HSCs, acceleration of neutrophil reconstitution after HSCT and mobilization of bone marrow HSCs into the peripheral blood. In recent years, several investigators have unraveled that G-CSF-mediated immune regulation including the switching T cell cytokine secretion profile to Th2 response, the altering adhesion activity of CD4+T cells to ICAM-1 and so on. Most of these cellular interactions of T cells are dependent on the integrin leukocyte function associated antigen-1 (LFA-1, αLβ2, CD11a/CD18) binding to intercellular adhesion molecule-1 (ICAM-1). Integrin αLβ2 adopts a bent, compact conformation in their low affinity state and an extended conformation in the high affinity state. The epitope of MEM148 is only expressed on the free β2 integrin subunit but is masked on LFA-1. The monoclonal antibody MEM148 reacts with an epitope exposed on free human CD18 chain as well as on high affinity state of LFA-1 that represents a reporter for αLβ2 active conformation. Our previous studies have demonstrated that rhG-CSF mobilization decreased the adhesion activity of CD4+T cells to ICAM-1, but did not find that rhG-CSF mobilization had effect on the level of CD11a expression. rhG-CSF maybe has effect on conformational change of LFA-1 on CD4+T cells. In order to explore the impact of rhG-CSF mobilization on LFA-1 conformation of CD4+ T cells, human CD4+T cells were isolated from peripheral blood mononuclear cells by positive selection with Miltenyi MACS. Isolated CD4+T cells were activated by OKT3+ICAM-1 and PMA+Ion separately. The slides with CD4+T cells were incubated with FITC-CD11a and FITC-MEM148 mAb, then were examined by using fluorescence microscope. The results show that the level of CD25, CD69 expression and MEM148 epitope exposure on activated CD4+T cells were significantly higher compared with unactivated CD4+T cells. rhG-CSF mobilization had no effect on the expression of CD11a, but could decrease expression of CD25, CD69 and exposure of MEM148 epitope on activated CD4+T cells. The percentage of LFA-1 polarized CD4+T cells before and after rhG-CSF mobilization was 85.32%, 61.86% respectively (p <0.01). No MEM148 epitope exposure on unactivated CD4+T cells was detected, but MEM148 epitope exposure on activated CD4+T cells before and after rhGCSF mobilization was 63.63%, 32.79% respectively (p <0.01). Overall, these data suggest that rhG-CSF can affect on the activation of CD4+T cells in mobilized hematopoietic stem cell allografts by altering the conformation of LFA-1.
Induced CD4+CD25+ FOXP-3+ and CD8+CD25+ FOXP-3+t Cells Exhibit a Concurrent Expression of Effector and Selective Suppressive Capacities
Authors: Avivi Irit , Stroopinsky Dina , Rowe Jacob , Avigan David , Katz Tami 
Published: October 2019

Abstract
Naturally occurring regulatory T cells (Tregs), identified as CD4+CD25+FoxP-3+ cells, are known to modulate immunologic tolerance and inhibit primary T cell activation, and therefore, may protect against graft-versus-host disease (GvHD) following allograft. However, having a non-selective suppressive effect, Tregs administration following transplantation may also result in delayed immunological reconstitution and diminished graft-versus-tumor effect, leading to an increased risk of infections and disease relapse. The generation of suppressive T cells that could selectively inhibit GvHD whilst preserving immunity against malignant cells and infection would promote significant contribution to GvHD prevention and therapy. The aim of this study was to investigate the characteristics of T cells undergoing allogeneic stimulation, looking for a subpopulation with a more specific inhibition of host immunity. A negative selection of PBMCs with anti-CD25 magnetic beads resulted in the production of CD25+ cells (-Naturally occurring) and CD25−PBMCs. CD25− PBMCs were than stimulated with allogeneic DC for 5–6. The resultant CD4+CD25+ or CD8+CD25+ populations were isolated with magnetic beads and investigated for immunophenotypical markers, cytokine expression profile (FACS), cell proliferation, inhibitory capacity and anti-viral response (INF-gamma). The current study has shown that stimulation of CD25− T cells with allogeneic peripheral blood mononuclear or dendritic cells (DCs) induces generation of a large fraction of CD4+CD25+ and CD8+CD25+ T-cells. This process was found to be more rapid and prominent following DCs stimulation. The resultant cells exhibit a mixed phenotype, demonstrating expression of the regulatory marker, FoxP3, as well as the inflammatory cytokines, IL-2 and IFNγ. Similar to naturally occurring Tregs, the induced CD4+CD25+ and CD8+CD25+ have markedly inhibited alloreactive T cell expansion. However, in contrast to naturally occurring Tregs, the induced populations, did not suppress proliferation against the cytomegalovirus (CMV) recombinant protein, suggesting their potential for more selective inhibition of host immunity.
Expanded naturally occurring CD4+CD25+T Induced CD4+CD25+T cells Induced CD8+CD25+T cells GITR + ++ ++ Immunophenotype CTLA-4 + ++ ++ FoxP-3 + ++ ++ Self-proliferation − − − Suppression rate Primary allogeneic stimuli + ++ ++ CMV recall antigen stimuli + − − Cytokine expression IL-2 − + + IFN-γ − ++ ++ Cytotoxic capacity (granzyme B) − − +
According to these data, CD25− derived CD4+CD25+FOXP-3+ and CD8+CD25+ FOXP-3, appear to have a dual expression of both effector and selective suppressive capacities, and therefore, might be potentially ideal for the use as post-transplant GvHD prophylaxis.
Only MHC-Identical Donor CD4+CD25+ Regulatory T Cells Convey Full Protection from Lethal Graft-Versus-Host Disease
Authors: Albrecht Julia , Doser Kristina , Andreesen Reinhard , Ermann Joerg , Edinger Matthias , Hoffmann Petra 
Published: October 2019

Abstract
Natural CD4+CD25+ regulatory T cells (Treg) contribute to tolerance induction after transplantation. We previously showed that the adoptive transfer of donor-derived Treg cells prevents lethal graft-versus-host disease (GVHD) after allogeneic bone marrow transplantation (BMT) in murine disease models. In contrast, host-type Treg cells failed to protect when co-transplanted under identical conditions. We now examined whether MHC compatibility between Treg cells and conventional CD25−CD4+ and CD8+ T cells (Tconv) is required for the suppression of alloresponses, or whether elimination of host-type Treg by allo-aggressive donor Tconv cells occurred. To address this issue, mixed lymphocyte reactions were performed in which CFSE-labelled responder T cells (Tresp), Treg cells and antigen presenting cells (APC) were systematically varied with regard to their MHC haplotype. When BALB/c (H-2d) Tresp cells were stimulated with mixed BALB/c and C57BL/6 (H-2b) APC, cultures contained 26.0 ± 3.1% and 86.2 ± 2.2% proliferating CD4+ and CD8+ T cells, respectively, on 6 d. In the presence of syngeneic BALB/c Treg cells, proliferation was decreased to 9.1 ± 4.7% and 25.1 ± 4.9% for CD4+ and CD8+ Tresp cells, respectively. In contrast, in cultures with allogeneic C57BL/6 Treg cells, proliferation remained at 22.1 ± 1.8% for CD4+ and 89.6 ± 0.4% for CD8+ Tresp cells. Comparable results were obtained with C57BL/6 Tresp cells after stimulation with F1 (C57BL/6 × BALB/c; H-2b/d) or 3rd party (DBA/1; H-2q) APC. Lack of suppression in co-cultures of MHC-mismatched Tresp and Treg cells was not caused by an early elimination of allogeneic Treg cells, as those were still detectable after 6 d of allostimulation. In corresponding in vivo studies, CB6F1 or DBA/1 recipients were protected from lethal GVHD only when Tconv and Treg cells were derived from MHC-identical donors, but not when they were from two MHC-disparate strains. Transplantation of 1 × 106 C57BL/6 Tconv cells resulted in 100% lethality of CB6F1 recipients by d56. When co-transplanted with 1 × 106 C57BL/6 Treg cells, all recipients survived for 100d, whereas only 40% survived after co-transfer of the same number of BALB/c Treg (n = 15; p = 0.004). Similarly, when 1 × 106 BALB/c Tconv cells were transplanted into CB6F1 recipients, all animals died from GVHD by d46. In contrast, all recipients of BALB/c Tconv and Treg cells (ratio1:1) survived for 100d, but only 10% of recipient mice survived after co-transfer of C57BL/6 Treg (n = 10; p < 0.001). Similar results were obtained after BALB/c and C57BL/6 T cell transfer into DBA/1 (3rd party) recipients. In conclusion, these data indicate that MHC-identity between Tconv and Treg cells is required for maximum suppression of an alloresponse and that Treg cells isolated from a 3rd party donor might not be suited for the prevention of GVHD after allogeneic BMT.
Reduction of Inhibitory Anti-FVIII Antibody Titer by Using a B Domain Variant FVIII/N6 cDNA for Nonviral Gene Therapy in Hemophilia a Mice
Authors: Jirovska Dominika , Ye Peiqing , Pipe Steven W. , Miao Carol H. 
Published: October 2019

Abstract
Due to the large size of FVIII, a B-domain deleted FVIII (BDD-FVIII) cDNA is usually used for developing gene therapy protocols for treating hemophilia A. Inefficient transcription of wild- type FVIII cDNA can be overcome by deletion of the heavily glycosylated B-domain encoding portion of the gene. BDD-FVIII is as clinically efficacious and not more immunogenic than full-length recombinant FVIII. More recently, it was demonstrated that a partial deletion of the B-domain leaving an N-terminal 226 amino acid stretch containing 6 putative asparagine-linked glycosylation sites intact (FVIII/N6) was able to increase in vitro and in vivo secretion of FVIII by 10–15 fold. We have inserted this B domain variant FVIII/N6 cDNA into our liver-specific gene expression vector. The resulting construct, FVIII/N6 plasmid was delivered into the hemophilia A mouse liver by the hydrodynamic method. In control mice treated with BDD-FVIII plasmid (n=5/group), FVIII expression dropped to undetectable levels at 2 weeks post injection and high-titer anti-FVIII antibodies were generated in all the plasmid-treated mice. However, in mice treated with FVIII/N6 plasmid (n=5/group), one out of five mice never developed inhibitory antibodies and still had some FVIII gene expression (~10%) at 8 weeks post gene transfer. Three FVIII/N6 plasmid-treated mice developed anti-FVIII antibodies with significantly reduced inhibitor titer and only one mouse developed high-titer inhibitory antibodies. The CD4+ T cells isolated from the spleen of mice injected with FVIII/N6 constructs proliferated less in response to FVIII stimulation than those from mice injected with BDD-FVIII. These results indicate that FVIII/N6 protein is less immunogenic than BDD-FVIII. Interestingly, both BDD-FVIII and FVIII/N6 constructs produced similar levels of FVIII gene expression (100–300%) initially following nonviral gene transfer. However this could be due to saturation of the ER to Golgi transport apparatus for FVIII by the initial high-level gene expression. Gene expression levels produced by using reduced dosages of BDD-FVIII and FVIII/N6 plasmids are currently being evaluated and compared. These findings suggest that use of a FVIII/N6 construct decreases transgene-specific immune responses following nonviral gene transfer and facilitates long-term gene expression.
THP-COP Regimen for the Treatment of Peripheral T Cell Lymphoma and Adult T Cell Leukemia/Lymphoma: A Multicenter Phase II Study
Authors: Takamatsu Yasushi , Suzumiya Junji , Utsunomiya Atae , Maeda Koichi , Matsuoka Hitoshi , Suzushima Hitoshi , Tsukada Junichi , Tamura Kazuo 
Published: October 2019

Abstract
Background: Pirarubicin (THP) is an anthracycline which was introduced to the Japanese market due to less cardiotoxicity compared to doxorubicin. The efficacy of THP-COP regimen (THP, cyclophosphamide (CPM), vincristine (VCR) and prednisolone (PSL)) was compared with CHOP in elderly patients with non- Hodgkin’s lymphoma in the previous Japanese randomized trial [Int J Hematol 81, 246–254, 2005]. The subset analysis showed peripheral T cell lymphoma (PTCL) had significantly better complete response (CR) rate with THP-COP than that of CHOP, whereas no such difference was observed in patients with B cell lymphoma.
Methods: We underwent a multicenter phase II study to investigate the efficacy of THP-COP as a first line treatment for PTCL and adult T cell leukemia/lymphoma (ATLL). THP 50 mg/m2, CPM 750 mg/m2, and VCR 1.4 mg/m2 were given intravenously on day 1. PSL 60 mg/m2 was administered orally on days 1–5. The treatment was repeated at a 21-day interval, and its efficacy and toxicity were investigated.
Results: Fifty-three patients aged from 38 to 83 (median 66) were entered into this study. Twenty-six patients were male and 27 female. Seventeen patients had PTCL including PTCL-U and AILT, and 36 ATLL. Stage IV disease was seen in 33, stage III in 15, and stage II in 5 patients. Thirty-four (64%) patients responded to THP-COP, including 17 (32%) CR and 17 (32%) partial response (PR). The median time to progression and overall survival were 9.5 and 17.5 months, respectively. Grade 3 to 4 neutropenia, anemia and thrombocytopenia occurred in 38 (72%), 18 (34%) and 31 (58%) patients, respectively. Twenty-seven (51%) patients developed febrile neutropenia, while grade 3 of nausea, anorexia, fatigue, arrhythmia, cardiac ischemia, sensory neuropathy, hypoalbuminemia, hyperbilirubinemia, and elevated serum AST/ALT were observed in 1 to 6 (2–11%) patients. No other adverse events greater than grade 3 were encountered.
Conclusion: THP-COP is an effective and well-tolerated treatment regimen in patients with PTCL and ATLL as a first-line therapy.