bluuebunny/crossref_metadata_2025_split huggingface dataset:
Identification of ADAMTS13 Epitopes Required for Binding to von Willebrand Factor Using Lambda Phage Surface Display.
Authors: Moriki Takanori , Maruyama Ichiro N. , Yamaguchi Yusuke , Igari Atsuko , Ikeda Yasuo , Murata Mitsuru 
Published: October 2019

Abstract
The metalloprotease ADAMTS13 cleaves multimeric von Willebrand factor (VWF) to regulate VWF-mediated thrombus formation. We planned to search core epitopes of ADAMTS13 that is required for its binding to VWF. We constructed a random cDNA fragment library expressing various peptides of ADAMTS13 on the surface of lambda phage and screened the library using immobilized VWF as a probe. After the first screening, the C-terminus of the spacer domain from Arg670 to Glu684 (termed as epitope-1) and the middle of the cysteine-rich domain from Arg484 to Arg507 (epitope-2) were determined as epitopes. When we added the synthetic epitope-1 peptide to the second screening, a new site, from Pro618 to Glu641 (epitope-3), was found in the middle of spacer domain. While the presence of synthetic epitope-2 peptide did not affect the subsequent screening, the presence of epitope-3 peptide enhanced the isolation of clones encoding epitope-1. These results suggest that ADAMTS13 epitopes-1, -2 and -3 may interact with each other for their binding to VWF. From screening in the presence of any combination or all of the three synthetic peptides, however, no new VWF binding site was uncovered. To examine the effect of divalent metal cations on the binding of ADAMTS13 epitopes to immobilized VWF, screening was carried out in the presence or absence of 5 mM of EDTA. No new epitope site was found. We next explored inhibitory effect of the synthetic epitope peptides on ADAMTS13 protease activity using recombinant ADAMTS13 and FRETS-VWF73 as a substrate. Synthetic epitopes-2 and -3 peptides markedly inhibited the cleavage of VWF by ADAMTS13, while the synthetic epitope-1 peptide did not as efficiently as epitopes-2 and -3. The stronger inhibitory effect of epitope-3 peptide than that of epitope-1 peptide was confirmed by SDS-agarose gel electrophoresis analysis of cleavage products of denatured multimeric VWF molecules by recombinant ADAMTS13. This was consistent with the dissociation constants for the three synthetic peptides with immobilized VWF determined by surface plasmon resonance, in which epitopes-2 and -3 have higher affinities for VWF than that of epitope-1. The results described above suggest that ADAMTS13 may initially bind to immobilized VWF through the sites of epitope-1 and epitope-2 with relatively weak affinity. The binding of epitope-1 to VWF may subsequently induce the conformational change of VWF, thereby exposing a binding site for epitope-3 for the efficient catalytic cleavage of VWF by ADAMTS13.
Prolonged Progression Free Survival Does Not Relate to Quality of Response to Treatment with Thalidomide in Patients with Relapsed Multiple Myeloma (MM).
Authors: Quach Hang , Prince Miles H. , Mileshkin Linda , Seymour John F. , Westerman David , Milner Alvin D. 
Published: October 2019

Abstract
Introduction: Quality of response to treatment in relapse/refractory MM, especially complete response (CR) or near complete response (nCR), has been reported to correlate with better PFS and OS in the past (Hussein MA, et al. Mayo Clin Proc2006;81:889–95). In this report, we provide an update on long-term survival from two multicentre phase II trials using thalidomide +/- IFNα2B (MM-thal) and combination celecoxib-thalidomide (Cel-thal) in relapsed or refractory MM, and identify predictors of progression-free survival (PFS) beyond 24 months (m).
Method: In 1998 and 2001, two prospective multicentre phase-II trials in relapsed or refractory MM were performed to assess the efficacy of thalidomide +/- IFNα2B (MM-thal), and combination celecoxib-thalidomide (Cel-thal), respectively. Both studies were previously reported (Blood2003;102:69; Clin Can Res2005;11:5504). The analysis of progression free survival (PFS) has been updated using the Kaplan-Meier method. Baseline characteristics were compared between patients having PFS ≥ 24m and < 24m using fisher’s exact test or the Cochran-Armitage test, to identify predictors of long-term disease control.
Result: Median follow up for MM-thal (n=75) and Cel-thal (n=66) trials were 73m and 47m respectively. Median PFS in the MM-thal trial was 5.5m, with estimated PFS of 9% at 3 years (95%, CI:5-18%), and 5% at 5 years (95%, CI:2-13%). In the Cel-thal trial, median PFS was 6.8m, with estimated PFS of 21% at 3 years (95% CI: 13-33%) and 16% at 5 years (95% CI:9-27%). Overall, 27 out of 141 patients (10 from MM-thal, 17 from Cel-thal) had PFS beyond 24m. The majority of these long term responders (70%) achieved only a PR as the best response to thalidomide-based treatment;15% achieved complete response (CR), and 15% had stable disease (SD). The most significant predictors for prolonged PFS of ≥24m was β2M ≥3mg/l (p<0.0005), stage ≥2 disease (p=0.001), and non-refractory disease to previous therapy (p=0.03). Bone marrow plasma cell infiltrate following thalidomide did not predict for outcome.
Conclusion: Thalidomide, and in particular combination celecoxib-thalidomide has substantial activity in relapsed MM with prolonged PFS beyond 24m in approximately 19% of patients. The strongest predictor of prolonged PFS is β2M. The depth of response to thalidomide had little influence on predicting remission duration.
Low Molecular Weight Heparin in Sufficient Dose of ≥70 IU/kg as an Effective and Safe Thromboprophylaxis in Patients with Newly Diagnosed Multiple Myeloma during Conventional VAD Induction Therapy.
Authors: Pour Ludek , Kessler Petr , Gregora Evzen , Zemanova Marketa , Penka Miroslav , Gumulec Jaromir , Adam Zdenek , Jaroslav Bacovsky , Frankova Hana , Havlikova Kamila , Lehanka Frantisek , Maisnar Vladimir , Meluzinova Ivana , Stavarova Yvetta , Straub Jan , Svobodnik Adam , Scudla Vlastimil , Sumna Elena , Walterova Lenka , Hajek Roman 
Published: October 2019

Abstract
Background: The patients with multiple myeloma (MM) have an increased risk of venous thromboembolism (VTE), particularly during the induction phase of therapy. Low molecular weight heparins (LMWH) seem to be useful for thromboprophylaxis in these patients. We analyzed prophylactic efficiency of LMWH based on dose either lower or higher than 70 IU/kg of body weight.
Patients and Methods: 223 patients with newly diagnosed MM were treated with induction chemotherapy consisting of vincristine, doxorubicin and dexamethasone (VAD) or vincristine, idarubicin and dexamethasone (VID) as a part of multicenter CMG 2002 trial. The first cohort of 137 patients received no systematic thromboprophylaxis. After a high incidence of thrombotic complications has been reported, the thromboprophylaxis with different doses of LMWHs was used in 86 consecutive patients for 4 months of induction therapy. Finally, 101 patients enrolled to the CMG 2002 trial in single centre were analyzed retrospectively, based on dose either lower or higher than 70 IU/kg of body weight. From this 101 patients 34% (34/101) received no systematic thromboprophylaxis, 38% (39/101) received LMWH (dalteparin) daily dose lower than 70 IU/kg, and 28% (28/101) received LMWHs daily dose higher than 70 IU/kg. The efficacy and safety of this treatment were analyzed. The chi-square test was used for statistical analysis.
Results: Incidence of VTE was significantly reduced in the group of 86 patients receiving the recommended thromboprophylaxis, as compared to 137 patients without prophylaxis (1.2% vs. 12.4%, p=0.003, risk reduction 11.25%). Single centre analysis showed that no VTE was developed in the group of 28 patients receiving more than 70 IU/kg of LMWH daily. It was significantly different from the 34 patients without prophylaxis (0% vs. 15%, p=0.002). The incidence of VTE in 39 patients with LMWHs daily dose lower than 70 IU/kg was reduced to 7.6%, which is still clinicaly significant as compared to cohort of 28 patients receiving more than 70 IU/kg of LMWH daily (0% vs. 7.6%, p=0.05). No case of major bleeding was developed in any group.
Conclusions: Thromboprophylaxis with LMWHs is effective in patients with newly diagnosed multiple myeloma during the induction chemotherapy only if the LMWH dose is sufficient. Our date shown that minimal sufficient dose seems to be more than 70 IU/kg of LMWH daily, as in this cohort of patients no case of VTE was developed.
Treatment of Patients with Multiple Myeloma Complicated by Renal Failure with Bortezomib - Based Regimens.
Authors: Roussou Maria , Kastritis Efstathios , Anagnostopoulos Athanasios , Psimenou Erasmia , Grapsa Irini , Matsouka Charis , Barmparousi Despina , Terpos Evangelos , Dimopoulos Meletios Athanasios 
Published: October 2019

Abstract
Introduction: Renal failure is a common feature of multiple myeloma and a major management problem. There is limited data regarding the reversibility of renal failure, the kinetics of serum creatinine and the safety of novel agents, such as bortezomib, when administered to newly diagnosed or relapsed/refractory patients with renal failure. The purpose of our analysis was to assess the frequency of renal failure improvement and kinetics of serum creatinine in patients who received bortezomib-based regimens.
Patients and methods: We evaluated 20 consecutive patients with newly diagnosed (n=7) or relapsed/refractory (n=13) multiple myeloma and renal failure, defined as a serum creatinine ≥ 2mg/dl. Patients’ median age was 66 years (range 43–88 years). Median serum creatinine was 3.8 mg/dl (range 2–11.9 mg/dl) and median creatinine clearance was 15.3 ml/min (range 6.4–33.3). Other features included hemoglobin <10gr/dl in 12 patients, platelets <100 × 109/l in 3 patients and elevated serum LDH in 9 patients. All patients received bortezomib plus dexamethasone alone or in combination with other agents, such as thalidomide, doxorubicin or melphalan. Reversibility of renal failure was defined as a sustained decrease of serum creatinine to <1.5 mg/dl and renal response was defined as ≥50% decrease of serum creatinine from its peak value.
Results: Reversal of renal failure was documented in 35% of all patients and the median time to reversal was 23 days. Moreover, 9 patients (45%) had 50% decrease in serum creatinine and the median time to decrease was 34 days. Some decrease of creatinine was documented in 88% of patients. Among four patients who were on renal dialysis, 2 became independent of this procedure after the second and the third cycle of treatment. The objective response rate was 61% and the median progression free survival for responders was 12 months. Toxicities were similar to those seen in myeloma patients without renal failure who were treated with bortezomib-based regimens. Grade 3–4 neutropenia and thrombocytopenia were seen in 28% and 22% of patients respectively. One patient died of infection and bortezomib had to be discontinued in 4 patients due to grade III neurotoxicity.
Conclusions: When bortezomib-based regimens are administered to myeloma patients with renal impairment their toxicity and efficacy are similar to those observed in patients with normal renal function. Moreover, these regimens are associated with rapid improvement of renal function in most patients and with reversal of renal failure in one-third of them.
PD-1 Expression Is Markedly Upregulated on Intratumoral CD4+ and CD8+ T Cells in Follicular Lymphoma and Is Associated with T-Cell Exhaustion.
Authors: Nattamai Durgha , Neelapu Sattva S. 
Published: October 2019

Abstract
Follicular lymphoma is one of the most immune-responsive of all human malignancies. However, immunoregulatory mechanisms in the tumor microenvironment may impair the efficacy of immunotherapies such as vaccines and adoptive T-cell therapy. The inhibitory receptor programmed death 1 (PD1), a negative regulator of activated T cells was recently shown to be upregulated on the surface of HIV-specific CD4+ and CD8+ T cells in humans and was associated with impaired T-cell function. Blockade of the immunoregulatory PD-1/PD-ligand 1 (PD-L1) pathway with antibodies against the PD-L1 augmented the function of HIV-specific CD4+ and CD8+ T cells (Day CL et al, Nature, 2006). To investigate the role of PD-1 in lymphoma, we examined PD-1 expression on peripheral blood mononuclear cells (PBMC) and intratumoral T cells in patients with follicular lymphoma prior to therapy. We observed that PD-1 expression is significantly upregulated on peripheral blood and intratumoral CD4+ and CD8+ T cells in patients with follicular lymphoma as compared with normal donor PBMC. Furthermore, PD-1 expression was significantly higher on intratumoral (mean 61%, range 34% to 86%) compared with peripheral blood CD4+ T cells (mean 25%, range 9 to 40%). Likewise, PD-1 expression was significantly higher on intratumoral (mean 44%, range 31% to 69%) compared with peripheral blood CD8+ T cells (mean 16%, range 9 to 31%). PD-1 expression on CD4+ and CD8+ T cells was associated with impaired type 1 cytokine production (IL-2, TNFa, and IFNg) and blockade of the PD-1/PD-ligand pathway with antibodies against PD-1 significantly enhanced T-cell function. These data indicate that the immunoregulatory PD-1/PD-ligand pathway is operative in patients with follicular lymphoma. Moreover, this pathway of reversible T-cell impairment provides a potential target for enhancing the function of exhausted T-cells in follicular lymphoma in combination with other immunomodulatory strategies such as vaccines and adoptive T-cell therapy.
Efficient Lysis of Myeloma Cells by CD16 Positive Effector Cells with Recombinant Single Chain Bispecific Antibodies Targeting HM1.24.
Authors: Peipp Matthias , Ehlert Claudia , Staudinger Matthias , Bruenke Joerg , Fey Georg , Gramatzki Martin , Repp Roland 
Published: October 2019

Abstract
Different therapeutic options are available for the treatment of multiple myeloma patients, but conventional chemotherapy often is not able to completely eradicate the tumor. After high dose chemotherapy, complete remission with only minimal residual disease could be achieved in many patients, but additional targeted strategies may help to eradicate residual cells and improve prognosis. Therapy with monoclonal antibodies is well established in CD20 positive B-cell lymphomas, but in contrast not many suitable target antigens are defined / expressed by multiple myeloma cells. HM1.24 a surface molecule expressed on terminally differentiated B-lineage cells represents a promising candidate antigen that is overexpressed on multiple myeloma cells. Here the development of a recombinant bispecific single chain Fv HM1.24 × CD16 antibody (tandem format) with novel features is presented. The HM1.24×CD16 bispecific antibody was expressed in 293T cells and purified to homogeneity by two-step affinity chromatography. Binding to HM1.24 and CD16 was demonstrated by immunofluorescence staining and flow cytometry with antigen positive and negative cells. The lytic activity of the bispecific HM1.24×CD16 scFv was evaluated in an antibody-dependent cellular cytotoxicity (ADCC) assay with different myeloma cell lines (RPMI 8226, INA-6, U266, JK6L) and primary patient derived cells as targets. Mononuclear cells (MNC), isolated from healthy donors served as effector cells. The bispecific HM1.24×CD16 scFv mediated efficient lysis of all tested cell lines at concentrations as low as 1 nM. In direct comparison to an HM1.24-IgG1 control molecule, the recombinant bispecific antibody demonstrated superior lytic activity at saturating concentrations and showed significant enhanced killing capacity. In conclusion, the recombinant bispecific HM1.24×CD16 retained its antigen specificity and demonstrated efficient lytic activity against patient-derived tumor cell lines and primary material. These results indicate that the bispecific antibody may be promising as a new therapeutic strategy in multiple myeloma.
Efficient Generation of Tumor-Specific, Cytotoxic T Cells by Genetic Transfer of allo-MHC.
Authors: Stronen Erlend , Abrahamsen Ingerid , Johansen Finn-Eirik , Gaudernack Gustav , Lund-Johansen Fridtjof , Olweus Johanna 
Published: October 2019

Abstract
Immune responses to established cancer are difficult to evoke by vaccination of patients due to various mechanisms of tolerance. In contrast, T cells in allogeneic stem cell grafts can contribute to cure of a number of hematological malignancies by the graft-versus-leukemia (GvL) effect. However, unknown T cell specificities causing life-threatening graft-versus-host disease (GvHD) limit the applicability of this treatment. Selection of GvL activity would increase the efficiency, safety and applicability of allogeneic stem cell transplantation. Here, we describe a novel method for overcoming tolerance by generation of allo-restricted, antigen-specific T cells. These cells specifically recognize and kill melanoma tumor cells presenting the melanoma-associated MART-1 in context of the prevalent major histocompatibility complex (MHC) class I molecule HLA-A*0201. HLA-A*0201-negative peripheral blood mononuclear cells were co-cultured with autologous, monocyte-derived dendritic cells transfected with HLA-A*0201 mRNA and subsequently pulsed with the peptide ELAGIGILTV from the melanoma-associated antigen MART-1. Antigen-specific T cells were detected in 8/10 donors on day 12 using peptide-MHC-pentamers (median 0.28% of CD8+ T cells, range 0-4.17%), and in all donors by day 19. When needed, cells were restimulated on day 12 with HLA-A*0201 mRNA-transfected and peptide pulsed autologous EBV-transformed lymphoblastoid cell lines (EBV-LCL). MART-1 pentamer+ T cells from five donors were sorted and expanded. The cytotoxic T cell lines (CTLs) responded with production of interferon-γ, degranulation and lysis of target cells when stimulated with peptide-pulsed EBV-LCL, even at low peptide concentrations (10−5 M). Similar responses were seen to melanoma cell lines expressing HLA-A*0201 and MART-1 (FM-57, Malme3M). In contrast, MART-1+ melanoma cells lacking HLA-A*0201 (Mel202) induced responses only when transfected with HLA-A*0201. Starting out with 0.5×106 pentamer+ T cells, a potential number of 3.0×109 pentamer+ T cells could be reached four weeks later, representing a 7800-fold expansion Our data show that allo-restricted T cells with high specificity for tumor-associated antigens can be rapidly generated, sorted and expanded to clinically applicable numbers.
Figure Figure
Therapeutic Effectiveness of Adaptive T-Cell Transfer for Minimal Residual Mantle Cell Lymphoma Following High Dose Therapy.
Authors: Joshi Shantaram S. , Munger Corey M. , Hegde Ganapati V. , Weisenburger Dennis D. , Vose Julie M. 
Published: October 2019

Abstract
Mantle cell lymphoma (MCL) is the most aggressive B-cell malignancy with an overall survival of less than four years. Although high-dose therapy followed by stem cell transplantation is effective in reducing the tumor burden and increasing the survival, the patients eventually relapse due to residual lymphoma. Therefore effective therapeutic modalities are needed to target residual MCL. In this study we have developed an immune-based approach to treat minimal residual MCL. HLA matched normal donor mononuclear cells were used to generate dendric cells in vitro. Following characterization, the dendritic cells were fused with the MCL cell line Granta 519 cells to create DC/MCL hybrids. The hybrids were then used to stimulate T cells from the same donor to generate MCL-specific cytotoxic T lymphocytes (CTL). The CTLs were tested for their MCL-specific cytotoxic abilities in vitro against MCL target cells and compared to control T cells or DC primed with MCL lysate or DCs transfected with MCL RNA. HLA matched human breast cancer cell line MDA-231 was used as an irrelevant tumor target. The in vitro results showed the highest MCL specific cytotoxic effects of CTLs generated using DC-MCL hybrids for immunostimulation. As a logical next step, in vivo therapeutic affects the MCL-specific CTLs were tested using a preclinical animal model. Four groups of NOD-SCID mice, with a minimum 10 mice per group, were transplanted with Grant-519 human MCL cell line intravenously. MCL bearing mice were treated with combination chemotherapy (CHOP)over three days followed by transplantation of 1 ×106 mononuclear cells. Ten days after, mice were transplanted with 3 ×106 adoptive T cells weekly for four weeks. The results of the in vivo studies showed that, adoptively transferred T cells that had been stimulated with DC/MCL hybrids were able to significantly inhibit tumor growth in the liver (p = 0.005), kidneys (p = 0.001), and lungs (p= 0.001) of mice with minimal residual MCL compared to control tumor bearing animals as determined by analyzing tumor burden in the organs using virtual images and Neuroinformatica software. When combined with CHOP chemotherapy, adoptively transferred T cells were able to significantly (P = 0.001) extend the survival of the mice by significantly further reducing the tumor burden as compared to mice only receiving the chemotherapy plus unmanipulated T cells. Specificity of the CTL mediated response was also confirmed by cytokine profiling in vitro and in vivo. In summary, we have clearly shown that antigen-primed DCs can be used to activate MCL-specific T cells which, when administered adoptively, are effective in treating minimal residual MCL. Thus these studies lay foundation for a PhaseI/II clinical trial.
(This study was supported by the Lymphoma Research Foundation, New York, NY).