bluuebunny/crossref_metadata_2025_split huggingface dataset:
Robust Expansion of Cytomegalovirus (CMV) Antigen-Specific CD4+ and CD8+ T Cells Using Gene-Modified Activated T Cells as Antigen Presenting Cells.
Authors: Melenhorst J. Joseph , Solomon Scott R. , Mielke Stephan , Hensel Nancy F. , Barrett Austin J. 
Published: April 2020

Abstract
CMV reactivation after stem cell transplantation can be treated with CMV-specific T cells but current in vitro techniques using dendritic cells as antigen-presenting cells are time-consuming and expensive. To simplify the production of clinical grade CMV specific T cells, we evaluated gene-modified activated T cells (T-APC) as a reliable and easily produced source of APC to boost CD4 and CD8 T cell responses against the immunodominant CMV antigen pp65. Peripheral blood mononuclear cells from CMV seropositive donors were activated for 2–3 days in complete medium (IMDM, AB serum, glutamine, and antibiotics) supplemented with 0.8 mg/ml phytohaemagglutinin and 100 IU IL-2/ml. The cells were transduced with Phoenix-A derived recombinant virus encoding pp65 in retronectin-coated 6-well plates, and further expanded in anti-CD3 plus CD28-coated flasks for 3–7 more days. Cultured cells expressed high levels of HLA-DR, and the costimulatory molecules CD80 and CD86. Autologous PBMC (0.5 – 1.0 x 106 cells) were stimulated with 106 irradiated (25 Gy) transduced T-APC in a 24-well plate. After 1–3 days IL-2 and IL-7 were added to a final concentration of 20 IU/ml and 10 ng/ml, respectively. Two weeks later the T cell lines were tested for antigen specificity using the flow cytometric intracellular detection of interferon-gamma following stimulation for 6 hours with a pp65 peptide library of 15-mers, overlapping by 11 amino acids. This technique induced a 135-fold (median; range, 20–120,000) and a 255-fold (median; range, 17–20,000) expansion of pp65-specific CD4 and CD8 responder cells, respectively, in 10/10 seropositive donors (figure). To further improve proliferation, CD25-expressing T regulatory cells were removed from the PBMC at the start of the culture by immunomagnetic depletion (Miltenyi). In 7/10 donors, CD25 depletion resulted in increased CD4 and/or CD8 responder numbers (p&gt;0.05; Mann Whitney paired t-test). Median increase in responder cell numbers was 4.25-fold (range, 1.4–6) for CD4+ T cells, and 4.2-fold (range, 3–7.5) for CD8+ T cells. These data indicate that T-APC efficiently boost pp65-specific CD4 and CD8 T cell numbers to clinically useful levels and that removal of CD25-expressing cells can further augment the total yield of antigen-specific T cells in most donors. The approach has the advantage of using a single leukocyte collection from the donor to generate large numbers of CMV-specific T cells within a total 3 week culture period using only one stimulation of antigen.
Fold-change in the total number of pp-65 specific CD4 and CD8 T cells from PBMC Fold-change in the total number of pp-65 specific CD4 and CD8 T cells from PBMC
The Immunomodulatory Drug Lenalidomide (CC5013; Revlimid), Enhances Antigen-Presenting Cell’s Function Leading to Effective Priming of CD4+ T-Lymphocytes.
Authors: Wang Hongwei , Naing Aung , Cheng Fengdong , Horna Pedro , Suarez Ildelfonso , Brayer Jason , List Alan F. , Sotomayor Eduardo M. 
Published: April 2020

Abstract
Professional antigen-presenting cells (APCs) play an important role in the initiation of antigen-specific T-cell responses. The demonstration that these cells are also required for the induction of T-cell tolerance, placed APCs at the crossroads of immune activation versus immune tolerance. Recent studies have demonstrated that the inflammatory status of the APC at the time of antigen presentation is the central determinant of T-cell priming versus T-cell tolerance. As such, therapeutic induction of inflammatory APCs might override immune tolerance and enhance the efficacy of immunotherapeutic strategies targeting hematologic tumors.
Lenalidomide (CC5013) is a thalidomide analogue with immunomodulatory properties. Phase I and Phase II clinical trials in patients with myelodysplastic syndrome (MDS) have shown high frequency of erythropoietic responses, particularly in patients with 5q31 deletion associated with emergence of polyclonal lymphoid infiltrate in responding patient bone marrows. This observation raised the question as to whether immunological mechanism(s) may mediate, at least in part, the beneficial effect of CC5013 in patients with MDS. To gain further insight into the effects of Lenalidomide on APC’s function and regulation of antigen-specific CD4+ T-cell responses, we treated peritoneal elicited macrophages (PEM) and bone marrow-derived dendritic cells (DCs) with escalating concentration of Lenalidomide in vitro. Enhanced expression of both B7.1 and B7.2 co-stimulatory molecules was observed in Lenalidomide-treated APCs relative to untreated APCs. No difference in the expression of MHC class II molecules or CD40 was detected. Assessment of cytokine production by ELISA showed that Lenalidomide-treated APCs produce higher levels of TNF-a, IL-6 and IL-10 in response to LPS stimulation as compared to untreated APCs. Next, we evaluated the ability of Lenalidomide-treated APCs to present cognate antigen to naïve and tolerant CD4+ T-cells specific for a MHC class II restricted epitope of influenza hemagglutinin (HA). We found that treatment of either PEM or DC with low doses of Lenalidomide (range: 1.5–12.5 uM) significantly enhanced their antigen-presenting capabilities leading to effective priming of naïve CD4+ T-cells confirmed by their increased production of IL-2 and IFN-gamma in response to cognate antigen. Taken together, our results shows that by inducing inflammatory APCs, Lenalidomide directs the outcome of antigen-specific T-cell responses. Furthermore, they have broadened the scope of this drug as a promising adjuvant in cancer immunotherapy.
Cell Based Assays Completed with the Mantle Cell Lymphoma Cell Lines Z138 and NCEB-1 Indicate That Combinations of Bortezomid and Flavopiridol Interact To Achieve Synergistic Activity.
Authors: Kapanen Anita , Tucker Catherine , Chikh Ghania , Bally Marcel , Klasa Richard 
Published: April 2020

Abstract
Mantle cell lymphoma (MCL) has the poorest long-term survival of all the lymphoma subtypes. CHOP-like regimens currently represent the standard of care for individuals with MCL, but traditional chemotherapeutic regimens do not elicit disease free survival times that are comparable to other sub classes of B-cell lymphomas. Median survival rate of a MCL patient still is approximately 3 years. Therefore new, targeted therapeutics are needed to improve the clinical outcome of MCL. The aim of the study was to identify and characterize combination effects achieved when flavopiridol, a Cyclin D1 inhibitor, and bortezomib, a proteosome inhibitor, are used in a combination setting together or with other agents contained in the CHOP regimen. Two MCL cell lines, Z138 and NCEB-1, were tested for cytotoxicity of drugs alone and in combinations, where alamar blue was used to assess cell viability. The resulting data were analyzed by using the median effect principle introduced by Chou and Talalay. In addition, mechanisms governing measured cytotoxic effects and combination effects were studied by DNA staining (propidium iodine), Caspase 3/7 activation and by western analysis of cyclinD1, NfkB, Bcl-xl, Bax and Bcl-2. The combination of bortezomib and flavopiridol was found to be synergistic in both cell lines studied. Synergistic interactions were dependent on drug-to-drug ratio. In NCEB-1, a ratio 1:12000 (bortezomib:flavopiridol) determined based on the IC90 of each of the individual agents showed synergy over a broad range of fa values (representing the fraction of affected cells). At ratios based on the IC10 and IC50 of the agents when used alone produced effects that were estimated to be antagonistic by the median effect principle. In the Z138 cell line, drug to drug ratios based on the individual agent IC50 and IC90 resulted in synergistic interactions. Consistent with the objective of identifying synergistic combinations, a clear dose reduction was observed to achieve a given therapeutic goal (e.g. 90% cell kill). 10-fold less bortezomib and 55-fold less flavopiridol would be required to achieve 90% cell death/cytostasis based on addition of the agents alone. Mechanistic studies showed that bortezomib in the combination and alone activated the caspase-dependent apoptotic pathway. The combination of bortezomib and flavopiridol for treatment of MCL would appear to an appropriate choice for clinical evaluation, however novel strategies must be developed in order to insure that combination effects observed in cell based screening assays are capture in patients. In this regard it is important to consider drug-drug ratio effects as well as combination engendered toxicities, both parameters that can be evaluated through careful pharmacodynamic assessments in well defined animal models of MCL.
Heat Shock Protein 90 (HSP90) Is Overexpressed in Primary and Cultured Hodgkin Lymphoma (HL) Cells: Role of Therapeutic Targeting Using the Small Molecule Inhibitor 17-AAG.
Authors: Georgakis Georgios V. , Li Yang , Rassidakis George Z. , Medeiros L. Jeffrey , Younes Anas 
Published: April 2020

Abstract
Conventional chemotherapy is the golden standard for therapy of Hodgkin Lymphoma (HL). Nevertheless, considerable toxicity and secondary malignancies indicate the need for targeted therapy that preferentially kills the malignant cells. The molecular chaperone heat shock protein 90 (HSP90) is expressed in all mammalian cells, but it is overexpressed in malignancy. 17-AAG, a small molecule inhibitor of HSP90, has been shown to induce apoptosis and cell cycle arrest in a variety of tumor types. In the present study we show that HSP90 is overexpressed in the primary Hodgkin and Reed-Sternberg (HRS) cells, as well as in HL derived cells lines. Inhibition of HSP90 17-AAG showed antiproliferative effect in HL derived cell lines in a dose dependent manner. Cell death was due to apoptosis, as determined by Annexin-V staining and FACS analysis. Apoptosis was mediated by the activation of the caspase pathway, especially by caspase 8, 9, and 3. Inhibition of caspase activity by the pancaspase inhibitor Z-VAD-FMK partially reversed the 17-AAG lethal effect. 17-AAG had no significant on the level of the antiapoptotic Bcl-2 family members or the cellular or X-Linked inhibitors of apoptosis. In contrast, there was considerable degradation of cFLIP. Moreover, 17-AAG treatment reduced the intracellular levels of molecules that have been shown to be of key importance in HRS cell survival and proliferation, including AKT and the phosphorylated ERK1/2, but with minimal change in total ERK1/2. Cell cycle arrest was observed at G0/G1 or at G2/M phase, and was mediated by reduction in the levels of MDM2, cyclin D1 with cdk4 and cdk6, and cyclin B1. The potential synergy of 17-AAG with conventional chemotherapy and anti-TRAIL death receptor monoclonal antibody, was explored by the simultaneous incubation of HL derived cells with both doxorubicin or antibodies against TRAIL receptors R1 and R2, respectively. The combination of 17-AAG with doxorubicin or anti-TRAIL antibodies was significantly more effective than either agent alone. Based on these data we are conducting a phase II study of 17-AAG in patients with relapsed classical HL.
Preclinical Anti-Tumor Efficacy of ABT-510 (Thrombospondin-1 Mimetic Peptide), a Novel Anti-Angiogenic Agent, in Hematological Malignancies.
Authors: Cikojevic Anica , Wang Eunice S. 
Published: April 2020

Abstract
Thrombospondin-1 (TSP-1) is a multi-functional 420 kDa matrix protein implicated in cancer cell adhesion, migration, and invasion and is a potent anti-angiogenic agent. TSP-1 expression is inversely correlated with vascularization and malignant progression of breast and lung cancers but has not been extensively investigated in hematological malignancies. We found low total mRNA levels of TSP-1 expression by RT-PCR in half (7 out of 14) of human cell lines representing lymphoma, leukemia, and myeloma. We also found lower total mRNA TSP-1 levels in two thirds (10 out of 15) of primary patient leukemia samples as compared to human umbilical vein endothelial cells (HUVEC). Treatment of human leukemia and lymphoma cell lines with recombinant TSP-1 (100 mg/ml) or ABT-510 (a novel synthetic peptide derivative of TSP-1) decreased tumor cell proliferation as measured by tritiated thymidine incorporation. Single agent ABT-510 treatment (total daily doses 50–120 mg/kg i.p. daily) resulted in statistically significant reduction in tumor volumes of subcutaneous SKI-DLBCL (diffuse large B cell lymphoma) xenografts. ABT-510 treatment (60 mg/kg IP bid) in disseminated SKI-DLBCL lymphoma mouse models resulted in increased long-term survival consistent with delayed disease progression. We then examined the effects of ABT-510 treatment in combination with anthracycline chemotherapy used in standard lymphoma regimens (doxorubicin) or another anti-angiogenic agent (anti-hVEGF antibody Avastin) in subcutaneous lymphoma xenografts. Our results suggest an additive effect with concomitant ABT-510 (60 mg/kg bid) and doxorubicin (3 mg/kg TIW) treatment, but no additive effect with concomitant ABT-510 (60 mg/kg bid) and Avastin (3 mg/kg TIW) treatment. Conclusion: Our data suggest that decreased levels of thrombospondin-1, a matrix protein with endogenous anti-angiogenic activity, play a role in the growth of human hematological malignancies. Based on these results, the TSP-1 synthetic peptide, ABT-510, represents a novel anti-angiogenic therapeutic strategy for patients with these diseases and warrants additional preclinical investigation in combination regimens. (Of note, a phase II trial of single agent ABT-510 in refractory/relapsed lymphoma patients has recently completed enrollment).
Nordic Mantle Cell Lymphoma (MCL) Project: Preemptive Rituximab Treatment of Molecular Relapse Following Autotransplant Can Reinduce Molecular Remission and Prolonged Disease-Free Survival.
Authors: Geisler Christian H. , Elonen Erkki , Kolstad Arne , Laurell Anna , Pedersen Lone B. , Andersen Niels S. , Jerkeman Mats , Nordstrom Marie , Lauritzen Grethe F. , Ralfkiaer Elisabeth , Akerman Maans , Sundstrom Christer , Langholm Ruth , Karjalainen-Lindsberg Marja-Liisa , Kuittinen Outti , Eriksson Mikael ,   Nordic Lymphoma Group
Published: April 2020

Abstract
The 2nd. Nordic Lymphoma Group mantle cell lymphoma (MCL2) protocol has demonstrated the importance of Ara-C and Rituximab in the induction chemotherapy and stem-cell mobilisation before high-dose therapy and autologous stem-cell transplant (1). By July 2005, 128 patients (83% stage IV) had completed protocol treatment consisting of 3 series of R-CHOP and 3 series of R-Ara-C, stem-cell harvest and high-dose therapy with BEAM/BEAC with ASCT. The 5-year failure-free and overall survival is 50% and 83% respectively, significantly higher than the historic control group of the Nordic MCL1 protocol with the same treatment without HD-Ara-C and Rituximab (P&lt;0.0001). Patients with a molecular marker (t(11;14) or clonal IgH rearrangement) identified at the time of diagnosis in bone marrow and blood, undergo regular molecular follow-up posttransplant,. Patients who turn PCR-positive or increase their qPCR signal, without clinical disease, are offered preemptive treatment with Rituximab 375 mg/m2 Wx4.
Of 75 patients with molecular markers who had completed treatment, 55 remain PCR-negative and 20 have become/remained PCR-pos. posttransplant. Clinical relapse ocurred significantly more often in the latter group (11 of 20) than in the PCR-neg. patients (4 of 55) (P&lt;0.0001) (Fig.1).
Ten of the 20 PCR-positive patient did not receive preemptive rituximab: five due to immediate clinical relapse, 2 due to stable qPCR signals, one due to protocol error and two await treatment. Of 10 patients who did receive preemptive rituximab 8 again became PCR-negative and 2 remain PCR-positive. Six of the 10 Rituximab treated patients remain in clinical and molecular remission 200–600 days after the Rituximab treatment (Fig. 2).
Conclusions:
In MCL, molecular relapse is a harbinger of imminent clinical relapse, whereas continuous molecular remission is associated with prolonged disease-free survival (89% at 4 years)
Rituximab preemptive treatment can reinduce molecular remission and may delay clinical relapse. Following molecular relapse, only Rituximab treated patients (6 of 8 evaluable) remain disease-free.
FIG. 1. NORDIC NCL-2 PROTOCOL: RELAPSE-FREE SURVIVAL ACC. TO MOLECULAR STATUS POSTTRANSPALNT FIG. 1. NORDIC NCL-2 PROTOCOL: RELAPSE-FREE SURVIVAL ACC. TO MOLECULAR STATUS POSTTRANSPALNT FIG. 2. NORDIC NCL-2 PROTOCOL: RELAPSE-FREE SURVIVAL FROM TIME OF MOLECULAR RELAPSE. FIG. 2. NORDIC NCL-2 PROTOCOL: RELAPSE-FREE SURVIVAL FROM TIME OF MOLECULAR RELAPSE.
Is First Line Single Agent Rituximab the Best Treatment for Indolent Non-Hodgkin’s Lymphoma? Update of a Multicentric Study Comparing Rituximab vs CNOP vs Rituximab Plus CNOP.
Authors: Rivas-Vera Silvia , Baez Enrique , Sobrevilla-Calvo Pedro , Baltazar Severiano , Tripp Francisco , Vela Jorge , Garces Oscar , Aguilar Luis , Ignacio Gregorio , Duque Jorge , Rodriguez Petty , Reyes Gustavo 
Published: April 2020

Abstract
Purpose: To evaluate efficacy, safety, Disease-Free Survival (DFS) and Overall Survival (OS) in patients with indolent non-Hodgkin’s lymphoma (NHL) treated with chemotherapy vs. immunotherapy vs immunochemotherapy as first-line therapy, an up-date report.
Methods: Patients with indolent NHL were randomized to receive: (A) Rituximab x 6/w, (B) CNOP (cyclophosphamide, mitoxantrone, vincristine and prednisone) x 6 or (C) R-CNOP x 6, at standard doses.
Results: 195 patients were included, 183 are evaluable for OS and toxicity (A:62, B:55 and C:66), 144 are evaluable for overall response rate (ORR) and DFS (A:53, B:41 and C:50).
Clinical characteristics: 89 male (45.6%), mean age 59±14 (±SD), 148 (75.9%) in stage (III/IV), without significant differences between groups. Overall Response Rate (CR+PR) was: A: 84.9%, B:83.4% and C:90% (P=0.545). Neutropenia grade 3/4 was more frequent in the chemotherapy groups: A: 4.8%, B: 23.6% and C:18.2% (P=0.001) as it was the infectious toxicity (grade 2/4): A:4.8%, B:5.5% and C:15.2% (P=0.07). DFS at 24 months was: A 68%, B:65% and C:70%, (P=0.93) and the OS was A:87%, B:84% and C:78%. P=0.89.
Conclusions: We did not find any important differences, between groups, regarding the Overall Response Rate, Disease Free Survival and Overall Survival at 24 months. However, single agent rituximab was better tolerated, with less toxicity in comparison with the chemotherapy containing groups. Based on these findings, it maybe reasonable to use immunotherapy only, as first-line therapy for patients with indolent NHL.
γδ T Cell-Mediated Immunotherapy of Malignancies with Zoledronate and IL-2.
Authors: Kunzmann Volker , Smetak Manfred , Kimmel Brigitte , Weissinger Florian , Weigang-Koehler Karin , Birkmann Josef , Einsele Hermann , Wilhelm Martin 
Published: April 2020

Abstract
Despite major advances in our understanding of the adaptive immunity towards tumors and the introduction of vaccine-based strategies, durable responses are rare and adaptive immunotherapeutic approaches are still not an established treatment modality. Several lines of evidence indicate that MHC-independent effector cells of the innate immune system such as natural killer (NK) cells or γδ T cells significantly contribute to the immune surveillance of tumors. As we have shown previously, aminobisphosphonates (ABP) such as pamidronate or zoledronate are potent γδ T cell stimulatory compounds by inducing secretion of proinflammatory cytokines (i.e. IFN-γ) and cell-mediated cytotoxicity against lymphoma and myeloma cells in vitro. The detection of ABP as γδ T cell stimulating drugs at pharmacologically achievable concentrations in humans opened the possibility to evaluate the consequences of selective γδ T cell stimulation in vivo. The concept of γδ T cell-mediated immunotherapy is currently validated in a Phase II clinical trial with zoledronate (4mg i.v., d 1) and low dose IL-2 (2 x 106 IU/m2 s.c., d 1–6) for patients with hematological (NHL, myeloma, AML) and non-hematological malignancies (renal cell carcinoma and malignant melanoma). The results of our first clinical pilot study with pamidronate/IL-2 in patients with lymphoid malignancies showed that selective activation and expansion of γδ T cells can be induced in vivo. However, 50% of patients with hematological malignancies failed to respond to pamidronate/IL-2 in vitro. Therefore, positive in vitro sensitivity testing was an essential inclusion criterion in this trial. Immunomonitoring of the first 12 patients included in the study showed that zoledronate/IL-2 is highly effective in activating and expanding γδ T cells in vivo (104 TCRδ2+/HLA-DR+ cells/μl (range 11-323) at day 8 of cycle 1 compared to 3 TCRδ2+/HLA-DR+ cells (range 0-15) before treatment). In addition, IFN-γ serum levels increased from 7 to 110 pg/ml (mean 44, n=6) at day 2 at cycle 1 (day 0: 2, range 0–5). So far, objective tumor responses were observed only in hematological malignancies (updated data will be presented). The application of zoledronate/IL-2 is generally well tolerated. In conclusion, effective γδ T cell activation/expansion can be achieved in vivo by the combination of zoledronate and low dose IL-2. Because of the potent anti-tumor effects of γδ T cells this strategy might be a new attractive immunotherapy approach for malignancies with preserved γδ T cell function.