bluuebunny/crossref_metadata_2025_split huggingface dataset:
The Role of Interleukin-15 on Natural Killer Cell Homeostasis and Activity in Patients with Acute Myeloid Leukemia
Authors: Szczepanski Mi roslaw J , Czystowska Malgorzata , Szajnik Marta E , Mandapathil Magis , Hilldorfer Benedict , Welsh Ann , Foon Kenneth , Whiteside Theresa L , Boyiadzis Michael 
Published: April 2020

Abstract
Interleukin-15 (IL-15) has been demonstrated to play a critical role in the regulation of natural killer (NK) cells. IL-15 induces the differentiation of NK cells from hematopoietic progenitors, stimulates the expansion of peripheral NK cells, and supports their survival. We investigated the role of IL-15 as a homeostatic regulator of NK cells in 29 patients diagnosed with acute myeloid leukemia (AML) and the potential role of IL-15 in enhancing the anti-tumor activity of NK cells in AML patients. The percentage of circulating NK cells was lower (p&lt;0.0001) in the AML patients (6%± 0.7, range 1–17%) compared to the NK cells of healthy donors (12%± 1, range 9–17 %). At diagnosis the mean level of IL-15 in patient plasma was 1.9 pg/ml (range 0.03–8.9) and increased (p &lt;0.02) to 5.2 pg/ml (range 0.06–13.4) after the completion of induction chemotherapy, when the NK levels had been reduced to zero cells/microliter. The mean level of IL-15 subsequently decreased to pre-treatment levels in the AML patients who achieved complete remission (mean 1.6 pg/ml, range 0.4–2.3). To assess effects of IL-15 on the NK cytotoxicity, we sorted NK cells from PBMC obtained from AML patients prior to treatment (at diagnosis) and cultured them in the presence of IL-15. Following IL-15 stimulation, a significant increase in NK-cell cytotoxicity against K562 targets and the patients’ autologous leukemic blasts was observed (p&lt;0.05) as was up-regulation in expression of the activating natural cytotoxicity receptors, NKp30 and NKp46 and the C-type lectin receptors NKG2D and NKG2C (p&lt;0.02–0.001). Addition of blocking antibodies to the activating receptors reduced NK-cell cytotoxicity. We determined that IL-15, a homeostatic NK-cell cytokine, increases after severe depletion of NK cells following intensive chemotherapy and this leads to increased NK-cell lytic activity in AML patients. These data suggest that modulation of IL-15 levels in AML could be therapeutically beneficial as IL-15 enhances NK-cell recovery following intensive chemotherapy and increases NK-cell anti-tumor activity.
Characteristics of Refractory and Relapsing Patients with Diffuse Large B-Cell Lymphoma
Authors: Coiffier Bertrand , Salles Gilles , Bosly Andre , Gaulard Philippe , Haioun Corinne , Casasnovas Olivier , Mounier Nicolas , Delfau Marie Helene , Sebban Catherine , Feugier Pierre , Recher Christian , Delarue Richard , Ketterer Nicolas , Tilly Herve , Gisselbrecht Christian 
Published: April 2020

Abstract
Even with the introduction of rituximab, some patients with lymphoma continue to relapse or progress during treatment. To better define these patients, we looked at all patients with aggressive lymphoma included in the GELA trials during the last 20 years or 4 generations of studies: LNH-87, -93, -98, and -03. Each study generation comprise several randomized studies according to different groups of patients, i.e., young or old, low or high risk. ACVBP, the high-dose regimen used by GELA since 1984 was usually one of the 2 arms, except in elderly patients. Rituximab was first introduced in the LNH-98.5 study and was part of nearly all arms in the LNH-03 study. A total of 7806 patients were included in this retrospective analysis, 3116 being treated with ACVBP and 4880 with other regimens. Two analyses were done: one for the 7198 patients treated without rituximab and one for the 608 patients treated with rituximab. Only patients included in a published study were included explaining the lower number for rituximab-treated patients, most of the LNH-03 studies being not yet published.
4 groups of patients were defined: refractory or non-responding pts (NR, progression during treatment); patients in PR at the end of treatment (persisting lymphoma cells or PET fixing tumor); relapsing pts with early relapses (ER, during the first year) and late relapses (LR, after one year). 7-year OS was 56% and 7-year PFS was 47.5%, meaning that only 8.5% of the patients were rescued by any treatment at time of progression. Identical results were found in all study generations with 2 groups of patients: those with PR or LR with 7-year OS at 38%, and those with NR or ER, 7-year OS at 12%. Therefore, all generations were grouped (see figure 1). Patients treated with rituximab had a gain of 9% in OS at 6 years (69% vs. 60%). However, they had the same pattern for progression: 6-year OS around 40% for PR and LR patients and around 20% for NR and ER patients. The difference between patients treated with or without rituximab being the percentage of patients in each group: 61% vs. 50% for those without progression and 16% vs. 29% for those with NR or ER. The IPI score does not allow the identification of these poor risk patients. A study is ongoing to characterize the NR+ER patients at diagnosis.
This analysis allowed recognizing 2 patterns of failing therapy. Patients with PR and LR have a disease sensible to chemotherapy while NR and ER pts have a disease not responding to treatment. These patients probably must be treated differently with the introduction of new therapeutic agents into the first line regimen.
Figure Figure
ADAMTS13 Gene Deletion Aggravates Ischemic Brain Damage
Authors: Nishio Kenji , Fujioka Masayuki , Hayakawa Kazuhide , Mishima Kenichi , Fujiwara Michihiro , Banno Fumiaki , Kokame Koichi , Miyata Toshiyuki , Shida Yasuaki , Sugimoto Mitsuhiko , Fukushima Hidetada , Okuchi Kazuo 
Published: April 2020

Abstract
Background: The proteolytic activity of human ADAMTS13 regulates the size of von Willebrand factor (VWF) multimers, controlling excessive platelet aggregation and preventing microvascular thrombus formation. Deficiency of ADAMTS13 causes thrombotic thrombocytopenic purpura (TTP) and patients with TTP often have neurological deficits such as stupor or coma. Therefore, ADAMTS13 appears necessary for vascular homeostasis in the brain and may also influence the response to brain injury during ischemic stroke.
Method and Result: We investigated the role of ADAMTS13 in a mouse middle cerebral arterial occlusion (MCAO) model of ischemia-reperfusion injury in the brain. We compared 24 wild type mice (WT) and 24 ADAMTS13 gene deleted mice (KO), which are healthy and fertile. All mice were males 6–8 weeks of age. Investigators were blinded to the genotype until all analyses were finished. We applied MCAO for 30 minutes followed by 23.5 hours of reperfusion. The cerebral blood flow (CBF) around the cortex of the ischemic region was measured by laser Doppler flowmetry for 60 minutes after MCAO. In both WT and KO mice, the CBF decreased to less than 20% of baseline during MCAO and returned to normal immediately after reperfusion. However, during the subsequent 30 min the CBF decreased to 34.6±5.8% of baseline for KO mice compared to 83.2±6.8% of baseline for WT mice (P &lt; 0.01) and remained significantly decreased at 24 hours, suggesting that ADAMTS13 deficiency promotes thrombosis after reperfusion injury. The infarction volumes of the brain were determined from the area not stained by 2,3,5,-triphenyltetrazolium chloride (TTC) 24 hours after MCAO. KO mice had significantly increased volume of brain infarction compared with WT (31.0±6.5mm3 vs 11.4±1.9 mm3, P &lt; 0.01). The degree of post-ischemic inflammation was estimated by Western blotting of plasma HMGB1 (high-mobility group box1) 24 hours after MCAO. Plasma HMGB1 was increased to 34.4 ± 5.3 ng/ml in KO mice, compared to 19.6 ± 3.5 ng/ml in WT mice (P &lt; 0.05). The KO and WT mice did not differ during the MCAO procedure in body temperature, survival (80% vs 85%) at 24 hours, prothrombin time, arterial pH, pO2 or pCO2.
Conclusion: ADAMTS13 deficiency aggravates the extent of persistent brain ischemia, infarct volume and inflammatory response after brief MCAO. Therefore, ADAMTS13 plays a neuroprotective role against ischemia-reperfusion injury.
Long Term Follow-up Analysis of HD9601 Trial Comparing ABVD Vs Stanford V Vs MOPPABVCAD in Patients with Newly Diagnosed Advanced Stage Hodgkin Lymphoma. A Study from the Intergruppo Italiano Linfomi (IIL)
Authors: Chisesi Teodoro , Bellei Monica , Luminari Stefano , Marcheselli Luigi , Levis Alessandro , Vitolo Umberto , Stelitano Caterina , Pavone Vincenzo , Merli Francesco , Liberati Marina , Baldini Luca , Bordonaro Roberto , Pesce Emanuela , Gobbi Paolo , Federico Massimo 
Published: April 2020

Abstract
PURPOSE: To provide long term follow-up results of HD9601 trial that compared ABVD vs MOPP-EBV-CAD (MEC) vs Stanford V (StV) regimens for the initial treatment of patients with advanced stage Hodgkin Lymphoma (HL)
PATIENTS AND Methods: Patients with stage IIB–III or IV were eligible for randomization among 6 cycles of ABVD, 6 cycles of MEC and 12 weeks of StV; treatment had to be consolidated with optional Involved Field radiotherapy on Bulky or slow responding sites. For the long term, follow-up analysis study database was updated with most recent follow-up data and with information on long-term toxicity.
Results: Between 1996 and 2000, 355 patients were registered into the trial and were randomly assigned to one of the three arms (ABVD 122 pts, MEC 106 pts, and StV 107 pts). Radiotherapy was administered to 62%, 47%, and 66% of cases in the 3 arms respectively. As previously described response rates at the end of treatment program were 89%, 94%, and 76% in the 3 arms, respectively. Initial results were published in 2005 with a median f-up of 61 months. Median follow-up of current analysis is 86 months. Compared with previous data we observed 2 additional relapses, both in StV arm. Moreover, we observed 7 additional deaths in patients who had achieved a CR after initial treatment. Overall, 20 patients died after achievement of CR with 7 additional events compared with our previous report (Gobbi et al. JCO 2007). Overall analyzing data by study arm we observed 4 (+3), 11 (+3), and 5 (+1) deaths in CR patients randomized toABVD, MEC, and StV, respectively. Additional deaths in CR from previous report were further analyzed and were caused by pulmonary embolism (3 cases: 2 ABVD, 1 MEC), sepsis (2 cases: 1 MEC, 1 StV), LMA (1 MEC), and second cancer NOS (1 ABVD). No additional significant longterm toxic event was recorded. Long-term analysis resulted in 8-yr OS of 88%, 84%, and 77% for ABVD, MEC, and StV, respectively without differences among study arms (P=0.337).
Conclusions: The long-term analysis of HD9601 trial confirmed the results of the initial analysis. Few additional events observed were mostly represented by deaths in CR patients but these events did not substantially modify survival rates of the three arms.
Stimulation of T Cells by Lenalidomide Involves Putative Lenalidomide Binding Proteins CD3-Epsilon-Associated Protein and GDP-Mannose Pyrophosphorylase a
Authors: Gandhi Ani ta K , Rogovitz Audrey , Lopez-Girona Antonia , Mendy Derek , Morrison Lisa , Xie Weilin , Schafer Peter H 
Published: April 2020

Abstract
Introduction: Lenalidomide is approved in the US for the treatment of transfusion-dependent patients with anemia due to Low- or Intermediate-1-risk myelodysplastic syndromes associated with a del (5q) cytogenetic abnormality, with or without additional cytogenetic abnormalities. Lenalidomide is also approved for use in the US and Europe in combination with dexamethasone in previously treated multiple myeloma patients. In vitro and ex vivo studies have shown that lenalidomide has a direct antiproliferative effect against tumor cells and induces antiangiogenesis. It has also been shown to have immunomodulatory activity, including co-stimulatory effects on T and NK cells. Using the yeast 3-hybrid system, based on a leukemia cDNA library and biotinylated lenalidomide analog, we identified 16 putative lenalidomide-binding proteins from 700,000 clones screened. These included CD3-epsilon-associated protein (CAST) and GDP-mannose pyrophosphorylase A (GMPPA). CAST binds to the T cell receptor and is also part of the RNA polymerase 1 complex. GMPPA is a nucleotidyl transferase that converts mannose-1-phosphate and GTP to GDP-mannose, which is involved in the production of N-linked oligosaccharides. To determine whether these proteins are required for lenalidomide-induced immunomodulatory activity, we evaluated the lenalidomide-induced upregulation of interleukin-2 (IL-2) production in primary T cells transfected with siRNA against CAST and GMPPA.
Methods: Primary human peripheral blood T cells were transfected with CAST or GMPPA siRNA, or mock siRNA as a control. Reduction of CAST and GMPPA expression was confirmed by qRT-PCR. After 24 hours, transfected cells were stimulated with anti-CD3 mAb in the presence or absence of 0.1 and 1 μM lenalidomide 48 hours. Cells were lysed after 48 hours, and IL-2 mRNA and mature protein levels were quantified using qPCR and ELISA, respectively.
Results: CAST gene expression in T cells was reduced by 60% (p&lt;0.05) in CAST siRNA-transfected cells. GMPPA gene expression was reduced by 66% (p&lt;0.05) in GMPPA siRNA-transfected T cells. Lenalidomide-induced IL-2 mRNA production was reduced from 9-fold in controls to 6-fold in CAST siRNA transfectants (p&lt;0.05) and from 15-fold in controls to 6-fold in GMPPA siRNA transfectants (p&lt;0.05). Lenalidomide-induced IL-2 protein production was reduced from 22-fold in controls down to 8-fold in CAST siRNA transfectants (p&lt;0.05) and from 5.6-fold in the controls down to 1.6-fold in GMPPA siRNA transfectants (p&lt;0.05).
Conclusion: These findings support the hypothesis that CAST and GMPPA play an important role in the lenalidomide IL-2 induction mechanism in primary T cells.
A Stapled p53 Helix Targets HDMX to Overcome Nutlin-3 Resistance and Reactivate the p53 Tumor Suppressor Pathway in Cancer
Authors: Bernal Federico , Wade Mark , Silverstein Amy M. , Verdine Gregory L. , Wahl Geoffrey M. , Walensky Loren D. 
Published: April 2020

Abstract
p53 is a transcription factor that induces cell cycle arrest or apoptosis in response to DNA damage and cellular stress, and thereby plays a critical role in protecting cells from malignant transformation. The E3 ubiquitin ligase HDM2 controls p53 levels through a direct binding interaction that neutralizes the transactivation activity of p53 and targets it for degradation via the ubiquitylation-proteasomal pathway. Whereas the HDM2-homologue HDMX lacks ubiquitin ligase function, it participates in regulating the p53 axis by heterodimerizing with HDM2 and sequestering p53 through protein interaction. Loss of p53 activity, either by deletion, mutation, or HDM2/HDMX overexpression, is the most common defect in human cancer. Tumors expressing wild type p53 are rendered vulnerable by pharmacologic approaches that stabilize and upregulate p53. In this context, HDM2 and HDMX have emerged as independent therapeutic targets for restoring p53 activity and resensitizing cancer cells to apoptosis in vitro and in vivo. The small molecule nutlin-3 is an effective antagonist of the p53-HDM2 interaction. However, several studies have demonstrated the inability of nutlin-3 to disrupt the p53-HDMX complex, rendering tumor cells that overexpress HDMX nutlin-3-resistant. We have previously described the synthesis and characterization of a hydrocarbon-stapled alpha-helical p53 peptide (SAH-p53-8) that binds HDM2 with low nanomolar affinity, targets HDM2 in situ, and reactivates the p53 tumor suppressor pathway in HDM2-overexpressing osteosarcoma cells. We now report that SAH-p53-8 binds HDMX with even higher affinity, co-immunoprecipitates with endogenous HDMX, and induces apoptosis and cell cycle arrest in nutlin-3-resistant cancer cells that overexpress HDMX. Thus, by inserting a chemical staple into a peptide fragment of the p53 transactivation domain, we have generated the first bifunctional inhibitor of HDM2 and HDMX, enabling the investigation and pharmacologic modulation of both targets in human cancer.
Development of a Novel Polyamide-Based Agent to Inhibit EVI1 Function
Authors: Sicot Géraldine , Vogel Marion , Zhang Yi , Harki Daniel A , Lezon-Geyda Kimberly , Dervan Peter , Perkins Archibald S. 
Published: April 2020

Abstract
The EVI1 gene at chromosome 3q26 is associated with acute myeloid leukemogenesis, due to both chromosomal rearrangement and to overexpression in the absence of rearrangement. Some rearrangements such as t(3;3) and inv(3) result in overexpression of EVI1 protein, while translocation t(3;21) yields an AML1-MDS1-EVI1 (AME) fusion protein. EVI1 possesses two zinc finger domains, an N-terminal domain with fingers 1–7, which binds to GACAAGATA, and a C-terminal domain (fingers 8–10) which binds GAAGATGAG. Inhibition of EVI1 function with a small molecule compound may provide a targeted therapy for EVI1-expressing leukemias. As a first step towards inhibiting the leukemogenic function of EVI1, we performed structure-function studies on both EVI1 and AME protein to determine what domains are critical for malignant transformation activity. Assays were
Rat1 fibroblasts in a soft agar colony forming assay for EVI1; primary bone marrow cells in a serial replating assay for AME.
Both assays revealed that mutation of arginine 205 in zinc finger 6 of EVI1, which completely abrogates sequencespecific DNA binding via the N-terminal zinc finger domain, resulted in complete loss of transforming activity; mutations in other domains, such as the C-terminal zinc finger domain, CtBP binding domain, and the domains of AML1 had less of an effect or no effect on transforming activity. In an effort to inhibit EVI1 leukemogenic function, we developed a polyamide, DH-IV-298, designed to block zinc fingers 1–7 binding to the GACAAGATA motif. DNAseI footprinting revealed a specific interaction between DH-IV-298 and the GACAAGATA motif; no significant interaction was observed elsewhere; a mismatch polyamide failed to footprint at equivalent concentrations; and DH-IV-298 failed to bind to a control DNA lacking the GACAAGATA motif. Electromobility shift assay showed that, at a 1:1 polyamide:DNA ratio, DH-IV-298 lowered EVI1:DNA affinity by over 98%, while mismatch was significantly less effective (74% reduction). To assess the effect of DH-IV-298 on EVI1 binding to DNA in vivo, we performed CAT reporter assays in a NIH-3T3-derived cell line with a chromosome-embedded tet-inducible EVI1-VP16 as well as a EVI1-responsive CAT reporter. Removal of tetracycline resulted in a four-fold increase in CAT activity that was completely blocked by DH-IV-298. The mismatch polyamide was significantly less effective than DH-IV-298. Further studies are being performed to assess the effect on endogenous gene expression, and on growth of leukemic cells that express EVI1. These studies provide evidence that a cell permeable small molecule compound may effectively block the activity of a leukemogenic transcription factor.
Development of mPF4/Heparin Antibodies in a Murine Platelet Activation Model
Authors: Suvarna Shayela , Qi Rui , Arepally Gowthami M 
Published: April 2020

Abstract
Heparin-Induced Thrombocytopenia (HIT) is an immune-mediated thrombotic disorder caused by antibodies that recognize complexes of Platelet Factor 4 (PF4) and heparin. We have previously shown that exogenous administration of murine PF4 (mPF4) and heparin (mP+H) daily for five days into mice leads to development of high titer PF4/heparin autoantibodies (anti-mP+H; Suvarna Blood, 2007). While this experimental model has been extremely useful for understanding various biological aspects of heparin sensitization, this murine model requires extremely high-doses of mP+H to elicit antibody production. To circumvent this non-physiologic approach, we studied effects of in vivo platelet activation in mice with or without concomitant heparin administration. After receiving approval from our Institutional Animal Care and Use Committee, we initially performed dose-finding studies of various platelet agonists {arachidonic acid (AA), ADP, collagen and PAR-4} to identify the maximal tolerated dose associated with in vivo platelet activation (assays of circulating PF4 levels) and associated with increased morbidity. Once optimal doses of platelet agonists were identified, we injected agonists in the presence or absence of heparin daily for five days and monitored for development of PF4/heparin antibodies as previously described (Suvarna, Blood, 2005). Of the various agonists tested, we noted development of anti-mP+H in mice injected with collagen (coll), but not in mice injected with AA, ADP and PAR-4. Moreover, anti-mP+H responses in mice injected with collagen showed heparin dependence, as shown in the Table below.
Mean A450nm Collagen (coll) alone (n=5) 0.04 ± 0.01 coll + 0.25 U/mL hep (n=5) 0.1 ± 0.04 coll + 0.5 U/mL hep (n=5) 0.1 ± 0.02 coll + 1 U/mL hep (n=5) 0.32 ± 0.27* (P&lt;0.05 v collagen alone)
Additional cohorts injected with coll + 1U/mL heparin showed significantly higher rates of seroconversion (mean A450nm= 0.25 ± 0.06, n=15) as compared to mice injected with coll alone (mean A450nm= 0.07 ± 0.02, n=15; p=0.007 for coll+hep v. coll alone, See Figure). In mice treated with coll+ 1U/mL Hep, anti-mP+H were specific for mP+H (mean A450nm= 0.358 ± 0.133) as compared to mPF4 alone (mean A450nm= 0.06±0.007). Anti-mP+H did not recognize rat albumin (mean A450nm= 0.015 ±0.006) or ELISA wells coated with PBS (mean A450nm=0.019 ±0.003). In summary, these studies indicate that anti-mP+H can be elicited in mice after platelet activation and heparin exposure. Further refinements of this model will be important in advancing our understanding the physiologic basis of heparin sensitization.
Figure Figure