bluuebunny/crossref_metadata_2025_split huggingface dataset:
HSV Amplicon Transduction Activates an Innate Immune Response in CLL B Cells;Implications for Immune Therapy and Vaccine Development.
Authors: Daubeuf Sandrine , Rosenblatt Joseph D. , Bowers William J. , Urbistondo Natasha T. , Federoff Howard J. , Tolba Khaled A. 
Published: April 2020

Abstract
Since gene therapy vectors are designed to shuttle therapeutic genes into various tissues, efficiency transduction is often viewed as the most critical measure of a vector’s clinical usefulness. However, other variables such as activation of the host’s innate immunity may play an equally important role shaping the clinical outcome. We have studied the application of helper-free HSV (hf-HSV) amplicon vectors to deliver costimulatory ligands into chronic lymphocytic leukemia (CLL) B cells for the purpose of enhancing their antigen presenting capacity. In this work, we examine the innate immune response of CLL B cells to transduction with helper-virus free HSV (hf-HSV) amplicon vectors. We screened culture supernatant of CLL B cells transduced with helper-free HSV amplicon for secretion of IL-6, TNF-α and interferon β by ELISA and observed significant induction of these cytokines upon hf-HSV transduction as compared to non-transduced and CLL cells transduced with helper virus-containing (H+-HSV) HSV amplicons. We also compared the effects of hf-HSV and H+-HSV amplicon vector transduction on surface expression of NKG2D ligand. CLL B cells transduced with hf-HSV but not H+-HSV amplicons upregulated the costimulatory NKG2D ligands MICA and MICB. To investigate mechanisms of HSV activation of innate immunity, we screened a panel of human TLR receptors for recognition of HSV amplicons using 293T cells transfected with individual human TLR receptors and assaying NF-κB driven luciferase activity as a read-out. We found that HSV amplicon activates TLR3 (receptor for double stranded RNA), TLR8 (receptor for single stranded RNA) and TLR9 (receptor for unmethylated CpG motifs). CLL B cells express predominantly TLR9 and to a lesser extent TLR8. TLR3, on the other hand is not expressed on resting B cells and is upregulated upon exposure to interferon β. In 293T cells lacking TLR receptors, viable hf-HSV but not UV-inactivated amplicons up-regulated surface expression of both MICA and MICB and activated interferon-β promoter driven luciferase, indicating the presence of a TLR-independent HSV activated innate response. To establish the presence of TLR-dependent NKG2D ligand induction, CLL B cells (expressing TLR7, 8 and 9) were incubated with their specific ligands (Loxaribine and CpG ODN motifs) for 24 hours and MICA and MICB expression confirmed by flow-cytometry. In summary, our data shows that hf-HSV amplicon vectors activate multiple aspects of innate immunity involving both TLR dependent and independent pathways, which translates into expression of cytokines and costimulatory ligands by transduced cells. The presence of helper virus on the other hand completely abolishes this response. In the setting of a vaccine development, the evoked innate response may prime the adaptive T cell response leading to improved clinical outcome.
Triterpenoid Methyl-CDDO Is a Potent Inducer of Apoptosis in CD34+ AML Progenitor Cells Via Activation of SAPK Pathways and Inhibition of MAPK Cascades.
Authors: Konopleva Marina , Ruvolo Peter , Contractor Rooha , Kurinna Svitlana , Shi Yue-Xi , McQueen Teresa , Milella Michele , Andreeff Michael 
Published: April 2020

Abstract
Outcome results in AML are a continued challenge for the development of novel therapeutic strategies. C-28 methyl ester of 2-cyano-3,12-dioxoolen-1,9-dien-28-oic acid, CDDO-Me, a novel triterpenoid, induces apoptosis in myeloid and lymphoid leukemic cell lines and in primary AML samples in sub-micromolar concentrations. We reported previously that CDDO-Me inhibits the activation of ERK1/2, blocks Bcl-2 phosphorylation, and promoted apoptosis in AML-derived U937 cells (Blood 2002, 99(1):326–35). Here, we examined the effects of CDDO-Me on CD34+ AML progenitor cells in vitro. Exposure to 1μM CDDO-Me induced apoptosis in all but one AML sample (46±4% annexin(+) CD34+ cells, n=20). To assess the anti-leukemia activity of CDDO-Me in vivo, scid mice intravenously injected with U937 cells were treated with liposomally-delivered CDDO-Me (20mg/kg/day IV every other day, starting at day 7, for a total of 9 injections). While CDDO-Me was not toxic to the mice, pathological and flow cytometry analysis revealed reduced (55–86%) leukemia burden in the bone marrow, liver, and spleen of mice. Since we had shown that CDDO-Me inhibits phosphorylation of pERK in U937 cells, a further goal of this study was to assess the role of ERK in CDDO-Me-induced cell death in primary AML samples. ERK was expressed and phosphorylated in all (n=15) patients’ samples studied. CDDO-Me inhibited ERK phosphorylation in 9 of 15 patient samples and promoted apoptosis. However, CDDO-Me still induced apoptosis in 5/6 samples that displayed no significant changes in pERK levels in response to the drug. This finding suggests that ERK is not the sole target of the compound. The stress-activated protein kinases JNK and p38 are related to ERK but in general activate pathways that are opposed to ERK. By Western blot analysis, CDDO-Me induced early (30 min) phosphorylation of JNK and p38, which preceded induction of cell death. Pre-treatment of U937 cells with JNK and p38 inhibitors SP600125 and SB203580 partially abrogated induction of apoptosis, while MEK inhibitor PD-98059 significantly enhanced cytotoxicity. CDDO-Me induced p38 phosphorylation in 5 of 6 primary AML samples tested. Collectively, these finding indicate that CDDO-Me markedly shifts signaling toward the JNK and p38 MAPK stress-related pathways and away from the cytoprotective MAPK cascade. In summary, the triterpenoid CDDO-Me is a potent inducer of apoptosis in primary AML including CD34+ AML progenitor cells. Induction of apoptosis is in part mediated via inhibition of ERK signaling combined with JNK/p38 activation. These studies in primary AML samples and the anti-leukemia activity of the compound in vivo suggest potential utility of CDDO-Me for the treatment of AML patients.
Generation of Human Anti-Leukemia CTL Vaccine Utilizing Dendritic Cell Hybrids.
Authors: Naparstek Elizabeth , Eshel Rinat , Vainas Oded , Ben-Yosef Rami , Shpringer Meirav 
Published: April 2020

Abstract
The therapeutic options for patients with hematological malignancies that relapse after autologous or allogeneic hematopoietic stem cell transplantation (SCT) are limited and their overall prognosis is poor. Therefore new therapeutic approaches such as cellular-based immunotherapies are being investigated. The efficacy of cytotoxic T lymphocytes (CTLs) or dendritic cell (DC) vaccines was already established in the treatment of post transplant CMV infections and in several solid tumors. This strategy however is less effective in hematological malignancies, where the tumor specific antigens are unknown and the cellular immunotherapy is based on the induction of graft-versus-leukemia (GVL) response by allogeneic, non-antigen specific, donor derived T lymphocytes (DLI). One approach to enhance immune recognition and targeting of the leukemic cells, and consequently augment their specific killing by CTLs, is through the use of dendritic cells. DCs are potent antigen presenting cells that can be manipulated to express tumor antigens by various strategies including fusion with tumor cells. Such hybrid cells expressing tumor antigens, presented in the context of MHC class I and II and co-stimulatory molecules of the DC partner, were shown to be effective in several malignancies.
Our aim was to generate human CTLs capable to mount a specific and potent in-vitro anti-leukemia activity, utilizing dendritic-leukemia cell hybrids. We have generated such hybrids by co-culture of immature human DCs with allogeneic (K562) or autologous leukemia cells in the presence of polyethylene glycol (PEG). After maturation, these dendritic-leukemia cell hybrids were utilized to generate specific CTLs, using autologous or allogeneic peripheral blood lymphocytes under IL-2 deprivation. Purified CD8+ cells were then further expanded and tested for their cytotoxicity against the target cells using Alamar Blue cytotoxicity assay.
Both allogeneic and autologous dendritic-leukemia cell hybrids were shown to stimulate a significant CTL responses compared to controls. Dendritic-K562 cell hybrids stimulated a CTL response of 40% lysis of the target K562 cells, while CTLs generated by co-culture with non-fused K562 cells had no detectable cytotoxic activity against their targets. Similarly, CTLs stimulated by dendritic-autologous leukemia cell hybrids showed 15% lysis of the leukemia target cells, compared to 1.5 and 1% lysis observed by CTLs generated by co-culture with autologous tumor or DCs alone. Our results show that in-vitro generation of hybrids of human dendritic and un-manipulated primary leukemia cells is feasible, and that such hybrid cells can elicit the generation of specific autologous or allogeneic anti-leukemia CTLs. Such CTLs may be superior to the currently used non-antigen-specific donor lymphocytes, and may be useful also in the autologous setting.
Selective Induction of Apoptosis in Acute Myelogenous Leukemia Stem Cells by the Novel Agent Parthenolide.
Authors: Guzman Monica L. , Karnischky Lilliana , Li Xiaojie , Neering Sarah J. , Rossi Randal M. , Jordan Craig T. 
Published: April 2020

Abstract
Acute myelogenous leukemia (AML) arises from a rare population of leukemic stem cells (LSCs), which are thought to perpetuate both de novo and relapsed disease. Because LSCs are often refractory to standard chemotherapy agents, it is crucial to investigate the ability of current and novel drugs to target this rare population of cells. Previously, we demonstrated that combined treatment with proteasome inhibitors and anthracyclines induced apoptosis in LSCs while sparing the normal hematopoietic stem cells (HSCs). Using this approach, death of leukemic cells was characterized by NF-kB inhibition and p53 activation. In the present study, we have extended this work by investigating parthenolide (PTL), a sesquiterpene lactone found in the herbal drug feverfew and that has been reported to inhibit NF-kB activation. Primary AML cells treated with increasing concentrations of PTL (1-10 microM) showed a rapid (18h) and dose dependent decrease in cell viability. The PTL concentration and its effect on induction of apoptosis directly correlated with decreased NF-kB binding activity and increased phosphorylation of p53. Importantly, apoptosis induction was also evident in the primitive LSC population, defined by the immunophenotypic profile CD34+, CD38− and CD123+. In contrast, PTL had little to no effect in normal HSCs obtained from either cord blood (CB) or bone marrow (BM). Interestingly, apoptosis induction upon PTL treatment was completely abolished upon treatment with the anti-oxidant, N-acetyl-cysteine (NAC), suggesting that PTL effects involve reactive oxygen species (ROS) induction. This was further corroborated using the fluorescent dyes DCFH-DA and mBBr to measure ROS and thiol content, respectively. When PTL effects were compared to the common chemotherapeutic drug cytosine arabinoside (Ara-C), we observed that PTL selectively targeted leukemic cells whereas Ara-C did not. Functional analyses using in vitro colony assays and the NOD/SCID mouse xenotransplant model were then employed to address whether the drugs used in this study affected stem cell potential. PTL treatment did not affect engraftment of normal cells while leukemic cell engraftment was dramatically impaired. Similarly, methylcellulose cultures showed a severe decrease on colony formation in PTL treated AML cells, while normal colonies were not impaired. Together, these data demonstrates that PTL is able to selectively target LSCs while sparing normal hematopoietic cells. The mechanism of leukemia-specific cell death appears to involve inhibition of NF-kB, activation of p53 and induction of oxidative stress.
Combination Chemoimmunotherapy Using Alemtuzumab, Fludarabine, Cyclophosphamide, and Doxorubicin (Campath-FCD) Is an Effective First-Line Regimen in Peripheral T-Cell Lymphomas.
Authors: Weidmann Eckhart , Hess Georg , Krause Stefan W. , Hebart Holger , Dreyling Martin , Subklewe Marion , Falge Christine , Chow Kai U. , Jaeger Elke , Mitrou Parisis S. 
Published: April 2020

Abstract
Peripheral (mature) T-cell lymphomas (PTCL) are represented by distinct lymphoma entities, many of which with a rather unfavourable clinical outcome as compared to B-cell lymphomas. Standard treatment regimens have not been established, except in primary cutaneous T-cell lymphomas (PCTCL). With regard to the unfavourable prognosis we designed a treatment protocol using chemoimmunotherapy in peripheral T-cell lymphomas except PCTCL and ALK-positive large cell anaplastic T-cell lymphomas, consisting of alemtuzumab 3, 10, 30, 30 mg, days 1–4, fludarabine (Flu) 25 mg/m2 days 2–4, cyclophosphamide (CP) 600 mg/m2 day 3, and doxorubicin (Dox) 50 mg/m2 day 4. Included were patients with primary diagnosis or with first or second relapse. Rationales for this regimen were proven efficacy of CP and Dox in T-cell lymphomas, high efficacy of Flu and CP (FC) in other lymphomas and the possible synergism of antibodies with cytotoxic drugs. So far, 23 patients have been included, 18 are evaluable for response and toxicity. Of the latter, 10 patients were diagnosed with PTCL-NOS, 5 with AILD, one with enteropathy-associated T-cell lymphoma, one with NK-cell lymphoma, and one with T-PLL. 9 patients were enrolled with primary diagnosis and 9 patients in relapse. The median age was 60 years (range 21–77); the median non-age adjusted IPI 2.5 (0–4), 2 in patients with primary diagnosis and 3 in patients with relapse. The overall response rate was 61% (11/18). In patients with primary diagnosis the CR rate was 78% (7/9), one patient achieved no change, and one patient died from treatment associated complications before response could be evaluated. All responding patients are in ongoing CR at 2+, 2+, 6+, 11+, 15+,16+, and 17+ months. In the group of relapsed patients one CR and 3 PR (44%) were observed. The main toxicity was leukocytopenia (81% grade III and IV of all evaluable treatment cycles), other grade III and IV toxicities included anemia (18%), thrombocytopenia (39%), infections (18%) nausea/emesis (9%), and allergic reactions (4%). Ten (56%) patients reactivated CMV without CMV-related disease and one patient with suspected CMV-peumonia. In conclusion, this is the first study demonstrating that alemtuzumab can be integrated into a triple-agent chemotherapy regimen. The combination is effective in the first-line treatment of peripheral T-cell lymphomas, however, regarding the general outcome a longer follow-up period of a larger patient population is required.
The Role of IL-12 and IL-23 in the Generation and Maintenance of Memory CD4+ T Cells.
Authors: Cleary Aileen M. , Lewis David B. 
Published: April 2020

Abstract
Human memory CD4+ T cells can be distinguished from antigenically-naïve CD4+ T cells based on their CD45RAlowCD45R0high and CD45RAhighCD45R0low surface phenotypes, respectively. Memory CD4+ T cells from adult peripheral blood can be further divided based on surface expression of the CCR7 chemokine receptor. Th1 memory CD4+ T cells that are CCR7high (putative central memory cells or Tcm) are expected to be preferentially targeted to peripheral lymph nodes where the ligands for CCR7 are expressed in high amounts. These cells have been reported to lack expression of the CCR3 and CCR5 chemokine receptors, which facilitate entry into inflamed tissues, and produce little or no interferon (IFN)-γ after stimulation via the αβ-TCR/CD3 complex. CD45RAlowCD45R0highCCR7low CD4+ T cells account for virtually all IFN-g production by human CD4 T cells after ab-TCR/CD3 stimulation using monoclonal antibodies, and for this reason were termed effector memory cells (Tem). These findings, as well as the observation of shorter telomere lengths for memory CD4+ T cells that are CCR7low compared to those that CCR7high suggest that the Tcm population may be an intermediate between naïve CD4+ T cells and Tem. It has recently been proposed that the level of signal strength and γc containing cytokines play a role in memory T cell generation. However, little is known whether IL-12 or IL-23 are necessary and for this differentiation and/or maintenance. Our laboratory has previously described a patient with IL-12Rβ1 deficiency, which ablates both IL-12 and IL-23 signaling. This patient had a deficiency in Tem number and function, unexpectedly suggesting that IL-12 and/or IL-23 may play a key role in this process. We therefore hypothesized that signaling through IL-12Rβ1 plays a key role in the latter stages of generation and/or maintenance of human memory CD4+ T cells. Preliminary data thus far show CCR7 expression to be slightly decreased on activated Tcm in response to incubation with IL-2 or IL-12 alone, and to a greater extent with IL-2 and IL-12 incubated together. In addition, spontaneous apoptosis of both Tcm and Tem is decreased upon incubation with IL-12. Taken together, these data suggest that IL-12 may play a role in both generation of Tem and maintenance of both Tcm and Tem.
Phenotype, Function, and Regulation of Migrating Hematopoietic Stem Cells.
Authors: Wagers Amy J. , Prohaska Susan S. , Passegue Emmanuelle , Price Jessica , Weissman Irving L. 
Published: April 2020

Abstract
Hematopoiesis in adult animals is maintained by a small population of clonogenic, multipotent hematopoietic stem cells (HSC), which maintain throughout life the capacity to self-renew and to differentiate to give rise to progeny cells that ultimately generate all lineages of mature blood cells. In adult mice and humans, the majority of HSC are found in the bone marrow (BM); however, HSC are also constitutively present at low levels in the circulation. The frequency of HSC in the blood can be significantly increased through the use of “mobilizing” agents, including cytotoxic drugs and/or cytokines, which often act both to drive HSC proliferation and to induce HSC migration from the BM into the bloodstream. Yet despite the increasingly common clinical exploitation of HSC in bone marrow and mobilized peripheral blood progenitor cell transplantation, both the evolutionary rationale and the molecular mechanisms that underlie the remarkable migratory capacity of HSC remain largely unknown. Therefore, to begin to elucidate the mechanisms and regulators of these events, we have used parabiotic and transplantation models to characterize normal blood-borne HSC. Our data clearly demonstrate that HSC are constitutively present in the blood of untreated mice and maintain a cell surface phenotype in the blood highly similar to their BM counterparts. Blood-borne HSC in normal mice can engraft both irradiated and non-irradiated BM niches, and subsequently are phenotypically and functionally indistinguishable from endogenous, host-type cells. These data suggest that BM homing of transplanted HSC in irradiated recipients and HSC mobilization in cytokine-treated animals likely makes use of pre-existing pathways that support the constitutive recirculation of these cells in normal animals. Finally, to extend these data and begin to uncover factors likely to play a role in stimulating HSC migration in both normal and mobilized mice, we have employed cDNA microarray technology to compare global gene expression profiles of normal and pre-migratory BM HSC, and have thus identified multiple candidate genes, including cell cycle regulators, signaling molecules, and transcription factors, that may be involved in HSC expansion or in HSC retention in and/or egress from the BM. Taken together, these findings provide significant insight into the dynamic nature and function of HSC, and may ultimately suggest novel and improved strategies for clinical hematopoietic cell transplantation.
Human Embryonic Stem Cell (hESC)-Derived Hematopoietic Elements Are Capable of Engrafting Primary as well as Secondary Fetal Sheep Recipients.
Authors: Narayan A. Daisy , Chase Jessica L. , Ersek Adel , Thomson James A. , Lewis Rachel L. , Kaufman Dan S. , Zanjani Esmail D. 
Published: April 2020

Abstract
We used transplantation into 10 and 20 pre-immune fetal sheep recipients (55–65 days-old, term: 145 days) to evaluate the in vivo potential of hematopoietic elements derived from hESC. The in utero human/sheep xenograft model has proven valuable in assessing the in vivo hematopoietic activity of stem cells from a variety of fetal and post-natal human sources. Five transplant groups were established. Non-differentiated hESC were injected in one group. In the second and third group, embroid bodies differentiated for 8 days were injected whole or CD34+ cells were selected for injection. In the fourth and fifth group, hESC were differentiated on S17 mouse stroma layer and injected whole or CD34+ cells were selected for injection. The animals were allowed to complete gestation and be born. Bone marrow and peripheral blood samples were taken periodically up to over 12 months after injection, and PCR and flowcytometry was used to determine the presence of human DNA/blood cells in these samples. A total of 30 animals were analyzed. One primary recipient that was positive for human hematopoietic activity was sacrificed and whole bone marrow cells were transplanted into a secondary recipient. We analyzed the secondary recipient at 9 months post-injection by PCR and found it to be positive for human DNA in its peripheral blood and bone marrow. This animal was further challenged with human GM-CSF and human hematopoietic activity was noted by flowcytometry analyses of bone marrow and peripheral blood samples. Further, CD34+ cells enriched from its bone marrow were cultured in methylcellulose and human colonies were identified by PCR. We therefore conclude that hESC are capable of generating hematopoietic cells that engraft in 1° sheep recipients. These cells also fulfill the criteria for long-term engrafting hematopoietic stem cells as demonstrated by engraftment and differentiation in the 20 recipient.