bluuebunny/crossref_metadata_2025_split huggingface dataset:
Engineered Oncogene Activation by Somatic Hypermutation Results in a Faithful Mouse Model of MGUS/Multiple Myeloma.
Authors: Robbiani Davide F. , Colon Kaity , Szabo Paul , Affer Maurizio , Nickerson Helen , Qi Chen-Feng , Morse Herbert C. , Chesi Marta , Bergsagel Peter Leif 
Published: October 2019

Abstract
The time in cellular differentiation at which the expression of an oncogene is dysregulated is thought to determine the phenotype of the resulting neoplasia. We tested this assumption in C57BL6/J transgenic mice expressing an HA-tagged c-myc oncogene in B cells under the control of kappa light chain regulatory elements. Two transgenes were engineered that differed by only one nucleotide.
The first set of mice carries a wild type HA-MYC transgene, develops pro-B cell tumors, and succumbs rapidly to pronounced lymphosplenomegaly.
In the second set of mice, a point mutation in the HA-tag portion of the transgene creates a stop codon that abrogates HA-MYC translation. This stop codon was engineered to be a hotspot for somatic hypermutation. Thus sporadically, in a germinal center B cell, somatic hypermutation may revert the stop codon, allowing translation of HA-MYC.
In two independently derived lines, mice spontaneously develop monoclonal gammopathies (50% incidence at 30 weeks (n=35), 80% at 40 weeks (n=35)). Serum protein electrophoresis detects M-spikes that increase in intensity over time, and 6 out of 7 cases tested so far were of IgG1 isotype. Unlike other mouse models of plasma cell neoplasia, no lymphosplenomegaly nor ascites were detected. In analogy to human multiple myeloma, that is a disease of isotype-switched and hypermutated cells homing to the bone, large populations of plasma cells were found in the bone marrow. Bone marrow (but not spleen) lysates reacted for HA and human MYC proteins by western blot and immunohistochemistry, thus indicating that reversion of the stop codon occurred. Remarkably, in two young mice, monoclonal spikes appeared 2 weeks after vaccination with NP-CGG. These spikes were sustained and the monoclonal protein was reactive to the NP antigen.
Together, comparison of the two sets of mice demonstrates that activation of the same transgene at distinct B cell developmental stages results in dramatically different tumor phenotypes. In addition, and opposite to most engineered mice, we have created a system where the oncogene is turned on sporadically, as occurs in the human disease. Furthermore, we have developed a model that faithfully reproduces important clinical aspects of monoclonal gammopathy of undetermined significance (MGUS) and its progression to multiple myeloma. This model will be useful to develop immunologic and chemoterapeutic approaches.
In Vitro Depletion of CD4+CD25+ T Regulatory (Treg) Cells with Denileukin Diftitox (DAB389IL-2) Increases T Cell Alloreactivity.
Authors: Lesnikova Marina , Nikitine Alla , Gass John , Nash Richard A. , Georges George E. 
Published: October 2019

Abstract
We hypothesize that immune tolerance after allogeneic hematopoietic cell transplantation is maintained by an active cellular mechanism that includes Treg CD4+CD25+ cells. In the dog model of mixed hematopoietic chimerism, adoptive immunotherapy with donor lymphocyte infusion (DLI) that successfully converted recipients to all-donor chimerism required the breaking of immune tolerance to target antigens. We reasoned that depletion of Treg cells might break tolerance and could improve the therapeutic efficacy of subsequent DLI. Denileukin diftitox (DAB389IL-2, Ontak) is a fusion protein consisting of the translocation and ADP-ribosylase domains of diphtheria toxin and interleukin 2. DAB389IL-2 binds to the high affinity IL-2 receptor (CD25, CD122 and CD132) expressed on the cell surface of activated T cells. Some Treg cells express the high affinity IL-2 receptor. Although DAB389IL-2 has been shown to deplete alloreactive T cells, we asked if DAB389IL-2 could deplete Treg cells without depleting alloreactive T cells in dog mixed lymphocyte cultures (MLC). First we asked if DAB389IL-2 could induce specific cell death of activated T cells. Five days after initiation of a one-way, dog leukocyte antigen (DLA)-mismatched MLC, DAB389IL-2 was added to the MLC for 24 hours and cells were pulsed with 3H-thymidine. At 10−11 and 10−10 M [DAB389IL-2] there was 34% and 92% inhibition of cell proliferation, respectively. To determine if DAB389IL-2 could deplete Treg cells, we established DLA-mismatched MLC (n=5) with cyclosporine (CSP) 400ng/mL (Martin, PJ et al. BBMT, 2004) and added DAB389IL-2 in half-log increments from 10−11 to 10−9.5 M on day 4 of MLC. On day 5, culture medium was changed to remove DAB389IL-2 and cells were analyzed for expression of CD4 and CD25. Flow cytometric analysis showed that CSP treated MLC had reduced CD4+CD25bright and CD4+CD25dim by 90 ± 3.3% and 64 ± 10%, respectively, compared to no CSP. DAB389IL-2 [10−11 M] treatment of MLC with CSP decreased CD4+CD25bright and dim populations 31 ± 12% and 30 ± 4%, respectively, compared to no DAB389IL-2. On day 10 of the culture, cells were tested in secondary (2°) MLC to determine the effect of depleting CD4+CD25+ cells with DAB389IL-2. 2° MLC was performed without the addition CSP or DAB389IL-2. After 3 days of 2° MLC, cells were assayed for proliferation. Compared to controls with no CSP or DAB389IL-2 treatment during 1° MLC, depletion of CD4+CD25+ cells with DAB389IL-2 increased 2° allo-specific proliferation 36.2 ± 26%. When CSP was present in the 1° MLC, DAB389IL-2 treatment on day 4 increased subsequent 2° MLC allo-specific T cell proliferation 86 ± 22%. These results are consistent with the hypothesis that DAB389IL-2 can efficiently eliminate Treg (CD4+CD25+) cells, particularly when Treg cells are generated in the presence of CSP.
Results of Unrelated Cord Blood Transplants in Fanconi Anemia.
Authors: Gluckman Eliane , Rocha Vanderson , Ionescu Irina , Pasquini Ricardo , Harris Richard , Wagner John , Kurtzberg Joanne , Duval Michel , Darbyshire P.J. , Locatelli Franco , Fernandez M.N. 
Published: October 2019

Abstract
Background: allogeneic hematopoietic stem cell transplant (HSC) is the only chance of cure in patients with Fanconi anemia (FA). Unrelated cord blood transplant is a possible alternative source of HSC. We report here a worldwide experience of unrelated cord blood transplant (UCBT) in a series of 72 patients with FA.
Patients and transplant characteristics : This is a retrospective multicenter analysis of 72 patients (62 children, 12 adults) with FA complicated by severe aplastic anemia or leukemia/MDS. Median age was 8 years ( 1–45), median follow-up after UCBT 23 months. The unrelated CB was HLA identical for A, B DRB1 in 9 cases, had 1 HLA difference in 29 cases and 2 or 3 HLA differences in 33 cases. The median number of nucleated cells (NC) infused was 4.4x107/kg, of CD34+ 2.1x105/kg. Conditioning varied: 24 patients received low dose cyclophosphamide with low dose TBI/TLI, 33 Fludarabine with cyclophosphamide and TBI or Busulfan, 55 patients received ATG. GVH prophylaxis consisted of cyclosporine alone (16) or with prednisone (37).
Results: CI of engraftment was 54%. In multivariate analysis: unfavourable factors were nb of HLA differences>2 (p .007) and NC infused <4.4x107/kg (p.04) CI of GVH> II was 32% and of cGVH 11%. Overall survival was 36%. In multivariate analysis, favourable factors were negative recipient CMV serology ( p.001), nb of NC infused> 4.4x107/kg (p.007) and use of Fludarabine (p.035). Causes of death were non engraftment :10 pts, GVH: 4 pts, infections: 15 pts and toxicity:15 pts. These results are quite similar to our current results of matched unrelated bone marrow transplant.
Conclusion: results of unrelated cord blood transplants could be improved by better patient and cord blood selection with units closely HLA matched and high number of cells, Conditioning with Fludarabine seems to improve results. In FA, use of cord blood might be beneficial because of the reduction of the risk of GVH which might decrease the risk of long term cancers.
Kit PY567- Directed Signals Are Important for Efficient Erythroid Progenitor Cell Proliferation and Survival, and Are Attenuated by Kit PY569.
Authors: Li Ke , Karur Vinit , Vincent Bethany , Curato Jennifer , Kapur Reuben , Besmer Peter , Wojchowski Don M. 
Published: October 2019

Abstract
Anemia incurred by mice with naturally occurring Kit mutations (eg, W41) clearly illustrates important contributions of SCF/Kit signaling to erythropoiesis. Specific cytoplasmic features of Kit and linked signaling events that support such contributions, however, are not well understood. To address this issue, the proliferative and survival signaling activities of a panel of Kit cytoplasmic PY mutants first were studied in SCF- and Epo- dependent G1E2 cells, and also in multi-potential SCF-dependent EML cells. Kit forms studied included PY567, PY569, PY567-plus-569, PY702, PY719, PY728 and PY745 phosphotyrosine add-back forms as well as a biologically inactive baseline mutant, Kit-F6 in which these six PY sites were mutated to phenylalanine. To by-pass endogenous Kit, cytoplasmic PY mutants were expressed from a MIEG3 vector as MCSF-R/Kit “MK” chimeras, and cells expressing matched and physiological levels of M/K chimeras were isolated by FACS. PY567 per se (ie, M/K PY567) selectively supported near wild-type proliferative and anti-apoptotic activities in both G1E2 and EML cells. By direct comparison, PY569 per se has diminished activity in G1E2 cells, and only baseline activity in EML cells. Moreover, the combined restoration of both PY567 and 569 uniformly attenuated the otherwise high proliferative and survival activities of PY567. These findings therefore indicate that Kit PY567 and PY569 do not necessarily exert overlapping roles, but that PY569 instead may preferentially link to negative effects and also act to attenuate key positive signals relayed via Kit PY567 (possibly in a lineage- restricted fashion). In keeping with recent studies in 32D cells, little if any bioactivities were conferred by PY702, 719, 728, or 745 per se. Mechanistically, PY567 also was observed to link with high efficiency to sustained Bcl-xl expression, and to be selectively efficient in Epo receptor co-signaling. While these findings predict that the mutation of Kit Y567 should perturb red cell development, erythropoiesis in Kit-F567 knock-in mice appears near-normal. However, primary erythroid progenitor cells isolated from the spleen of Kit F567 mice at 50hr post- phenylhydrazine injection proved to be markedly deficient in SCF responsiveness. In addition, a >5-fold decrease in SCF and Epo co-signaling was reproducibly observed for Kit-F567 progenitor cell preparations. These novel findings therefore lend new evidence to the case that PY567, in fact, contributes in important ways to Kit’s erythropoietic biofunction.
Signaling by the Granulocyte-Colony Stimulating Factor Receptor Involves Jak2-Dependent Phosphorylation of Gab2.
Authors: Wang Lin , Xue Jia , Corey Seth J. , Robinson Lisa J. 
Published: October 2019

Abstract
Granulocyte colony stimulating factor (G-CSF) is the major cytokine involved in neutrophil production. G-CSF has pleiotropic effects on myeloid cells, initially stimulating proliferation but later promoting differentiation. The specific signaling pathways that mediate the diverse effects of G-CSF remain incompletely understood. Recently, the scaffolding molecule Grb2-associated binder protein 2 (Gab2) was shown to play an important role in G-CSF induced myeloid differentiation (Zhu et al. Blood 2004). Ligand stimulation of the G-CSF receptor results in the rapid phosphorylation of Gab2, but the identity of the responsible kinases and the molecular events dependent on Gab2 phosphorylation remain unclear. Because Janus kinases (Jaks) play a central role in G-CSF signaling, we investigated the involvement of Jaks in G-CSF-stimulated Gab2 phosphorylation using the hematologic DT40 cell line stably transduced with the human G-CSF receptor (DT40GR). Antisense Jak1 and Jak2 constructs expressed in DT40GR cells each produced a marked reduction in their target Jak protein, but only antisense Jak2 reduced G-CSF-stimulated Gab2 phosphorylation. To determine whether Gab2 phosphorylation required Jak2 kinase activity, dominant negative Jak2 mutants lacking catalytic activity were expressed in the DT40GR cells. Expression of dominant negative Jak2 inhibited Gab2 phosphorylation in response to G-CSF. Similarly, treatment with the Jak2-selective kinase inhibitor AG490 markedly reduced G-CSF-dependent Gab2 phosphorylation. Co-immunoprecipitation studies further demonstrated a G-CSF- and Gab2 phosphorylation-dependent association of Jak2 with Gab2 in vivo, which was detectable by 30 seconds after G-CSF stimulation. To determine whether Gab2 was a direct substrate of Jak2, we performed in vitro phosphorylation studies using Gab2-GST fusion protein substrates. Jak2 immunoprecipitated from G-CSF-stimulated cells, but not from control cells, phosphorylated the Gab2 fusion protein. To identify potential Jak2 tyrosine phosphorylation sites in Gab2, we used site-directed mutagenesis to produce three Gab2 tyrosine mutants. Tyrosines 409, 452, and 476 were each replaced by phenylalanine (Y409F, Y452F, and Y476F). The Y452F and Y476F mutations of Gab2 each inhibited G-CSF-stimulated Jak2-dependent phosphorylation of Gab2, both in stably-transfected DT40GR cells and in transiently-transfected 293 cells also transduced with the G-CSF receptor. In contrast, G-CSF-stimulated Gab2 phosphorylation appeared unaffected by the Y409F mutation. We also evaluated downstream events in G-CSF signaling in cells expressing these Gab2 tyrosine- mutants. Akt and Erk phosphorylation following G-CSF stimulation was inhibited by both the Y452F and Y476F Gab2 mutations, but was unaffected by the Y409F mutation. These results suggest that Jak2 may mediate G-CSF differentiation signals through Stat-independent mechanisms.
Pilot Phase Results of the Human Proteome Organization Plasma Proteome Project.
Authors: States David J. , Adamski Marcin , Menon Rajasree , Hanash Samir , Omenn Gilbert S. 
Published: October 2019

Abstract
The protein components of blood play essential roles in coagulation, inflammation, immunity, growth and development. Recognizing the importance of blood proteins in health and disease, the Human Proteome Organization (http://www.hupo.org) chose the Plasma Proteome Project (PPP) as the first tissue specific analysis to be implemented. The PPP is a collaboration of more than 30 laboratories and research groups worldwide. The June 2004 PPP Jamboree held in Ann Arbor, MI marked the completion of the pilot phase of the PPP with more than 50,000 peptide identifications reported leading to a high confidence consensus list of about 2,300 human blood proteins. Important contributions of the pilot phase of the PPP include the identification of over 2 thousands human blood proteins, comparative assessment of proteomic analysis of serum and plasma prepared using citrate, EDTA and heparin anticoagulants samples prepared from a single well defined donor pool; assessment of abundant protein depletion technologies to enhance the dynamic range of proteome analysis; comparative assessment of different protein and peptide fractionation technologies; comparison of MALDI peptide fingerprinting, liquid chromatography electrospray tandem mass spectroscopy, direct MS/SELDI, and accurate mass tag strategies; correlation of protein abundance with success in mass spectroscopy identification; comparison of Sequest, Mascot, SpectrumMill and X!Tandem software for mass spectroscopy data analysis; comparison of NCBI NR, IPI and full 6-frame genome translation databases; establishing quality measures for the different software packages; establishing quality assurance and data validation protocols for large multi-center proteomics studies. These results demonstrate the potential for large scale proteome analysis as well as highlighting important challenges that lie ahead.
Economic Analysis of Prophylactic Pegfilgrastim in Cancer Patients Receiving Chemotherapy.
Authors: Eldar-Lissai Adi , Cosler Leon E. , Lyman Gary H. 
Published: October 2019

Abstract
Introduction: Neutropenia and its complications are major dose-limiting toxicities of systemic cancer chemotherapy. Febrile neutropenia (FN) generally prompts immediate hospitalization for evaluation and empiric broad-spectrum antibiotics adding to the cost of cancer care. Randomized controlled trials (RCTs) have demonstrated that prophylactic recombinant granulocyte colony-stimulating factor (rG-CSF; filgrastim) is capable of reducing the risk of FN, infection related mortality (IRM) and duration of hospitalization associated with FN. In the absence of compelling clinical indications, current guidelines support the use of prophylactic G-CSF only when using regimens associated with FN risk of 40% or higher. While recent RCTs have demonstrated that a long acting, pegylated form (pegfilgrastim) is at least as effective and safe as filgrastim, its economic impact has received little study.
Methods: An economic analysis was conducted comparing prophylactic pegfilgrastim to no prophylactic growth factor in patients receiving systemic chemotherapy. Direct medical cost estimates (US$ 2002) were obtained from the University HealthSystem Consortium and Marketscan databases. Pegfilgrastim costs were based on the average wholesale Red Book© prices. Risk and efficacy estimates were based on recent RCTs and meta-analysis of prophylactic rG-CSF. Sensitivity analyses and estimation of cost neutral thresholds for the range of feasible values of all variables were conducted based on cost-minimization.
Results: Under baseline conditions (FN= 17%; RRR=0.941), an incremental cost savings with pegfilgrastim of $428 was estimated. At baseline, a cost neutral threshold for FN risk of 14.6% was estimated with an expected cost for either strategy of $2721. Sensitivity analysis demonstrates a baseline threshold for RRR of 0.74 with lower rates of RRR supported by increasing hospital cost or length of stay, increasing infection-related mortality or incorporation of indirect costs. Threshold estimates for FN risk were robust over all possible values assumed for RRR of infection related mortality. The FN risk threshold falls below 10% when hospitalization cost per day exceeds $3000. Further results, including a Monte-Carlo simulation will be presented.
Conclusions: Incorporation of recent RCTs and cost data into clinically relevant models demonstrates that pegfilgrastim is cost saving at levels of FN risk associated with common chemotherapy regimens (< 20%). Primary prophylaxis with pegfilgrastim should be considered in patients being treated for curative intent with moderately myelosuppressive regimens.
Impact of Immunosuppressive Drugs on Adaptive and Innate Immune Responses to Aspergillus fumigatus Antigens.
Authors: Hebart Holger , Mickan Andreas , Haddad Ziad , Loeffler Juergen , Latge Jean-Paul , Einsele Hermann 
Published: October 2019

Abstract
Appropriate activation of the innate and adaptive immune system is crucial for the successful control of invasive aspergillosis (IA). Acute and chronic graft-versus-host disease as well as corticosteroids were described as major risk factors for the development of IA. In this study, we assessed the impact of immunosuppressive agents (dexamethasone, rapamycin, Cyclosporin A, FK506) on the A. fumigatus induced activation of monocyte-derived immature dendritic cells (iDC) and A. fumigatus-specific T-cell responses in well established cell culture models.
Immature DCs were found to be activated and to differentiate into mature DCs in response to A. fumigatus antigens. The upregulation of CD86 was inhibited by dexamethasone (D) in 3 out of 3 experiments, and of CD40 and CD80 in 2/3. CSA and FK506 had a variable impact on the upregulation of CD86, but not on CD40 and CD80, whereas the expression of co-stimulatory molecules was found unchanged upon incubation with rapamycin.
Autologous DCs were found to restore A. fumigatus-specific T-cell responses. T-cell proliferation to A. fumigatus hyphae and a cellular extract of the culture filtrate were found to be strongly inhibited by rapamycin and dexamethasone (n=3), whereas the effect of CSA and FK506 (n=3) at the concentrations analysed was variable. The release of IFN-g in culture supernatants upon stimulation with A. fumigatus antigens was strongly reduced in the presence of rapamycin (n=3), whereas the release of IL-4 was found to be increased in the majority of experiments (n=3). Comparable results were observed upon stimulation with tetanus toxoid and a CMV lysate (n=3). These data indicate, that A. fumigatus-spec. T-cell responses may be directed towards a TH2 phenotype in the presence of immunosuppressive agents.
In summary, immunosuppressive agents were found to exert differential effects on adaptive and innate immune responses directed against A. fumigatus. Whereas dexamethasone was found to modulate the expression of co-stimulatory molecules on A. fumigatus activated iDCs and to suppress A. fumigatus-specific lymphoproliferation, rapamycin exerted only minor effects on DC-activation but had a strong impact on A. fumigatus-induced T-cell responses. These results may help to tailor immunosuppressive regimens in patients at high risk for invasive aspergillosis.