bluuebunny/crossref_metadata_2025_split huggingface dataset:
Activated T Cell-Mediated Immunotherapy with a Chimeric Receptor Against CD38 in B-Cell Malignancies.
Authors: Mihara Keichiro , Yanagihara Kazuyoshi , Takigahira Misato , Ochiya Takahiro , Imai Chihaya , Kitanaka Akira , Takihara Yoshihiro , Kimura Akiro 
Published: April 2020

Abstract
CD38 is expected to be one of the most useful molecular targets on the surface of malignant B-cells. T cell-mediated immunotherapy with a chimeric receptor could provide a powerful tool for treating cancer. However, since CD38 is also expressed on effector cells such as activated T cells, NK cells, and monocytes, these cells with the chimeric receptor could be eliminated by autologous cytotoxicity through the interaction with the antigen. In this study, we developed a novel methodology for enhancing the survival and clonal expansion of T lymphocytes expressing an anti-CD38 chimeric receptor. Hut78 T cells, which express very little CD38, retrovirally transduced with the anti-CD38 chimeric receptor showed powerful cytotoxic activity against B cell lines expressing CD38, such as HT (lymphoma), RPMI8226 (myeloma), 380 (ALL-Ph1−) and OP-1 (ALL-Ph1+) cells (mean specific cytotoxicity was 97.94% ± 0.31% after four days of culture in vitro). However, in activated human T cells and Jurkat cells constitutively expressing CD38, the recovery rate of cells transduced with the chimeric receptor was extremely low, because the cells eradicated each other and/or themselves by inducing apoptosis. To block the interaction of the anti-CD38 chimeric receptor with CD38 antigen, we incubated activated T cells and Jurkat cells in medium supplemented with an anti-CD38 antibody before the transduction. The number of viable cells harvested after the transduction was dramatically increased by the antibody in a dose-dependent manner. Using this method, we prepared human peripheral T cells bearing the chimeric receptor and injected them into NOD/SCID mice, which were transplanted with HT cells labeled with luciferase. Lucuferase activity was not detectable in 13 days in five of six mice with T cells transduced with the chimeric receptor. In contrast, the activity had a rapid and steady increase in all of the mice injected with vector-transduced T cells. These results clearly showed that even though human peripheral T cells express any molecule on their surface, an antibody could protect T cells transduced with a chimeric receptor-containing vector from cytolysis, and apoptosis. These findings may provide us with a powerful tool for improving T cell-mediated targeting therapy.
Pharmacokinetics of Anti-CD22 Antibody Conjugates with Uncleavable and Cleavable Linkers in Rats.
Authors: Xie Dong , Deng Rong , Baudys Jakub , Chan Pam , Dere Randy , Ebens Allen , Fielder Paul , Fuji Reina , Gray Alane , Hung Catherine , Iyer Suhas , McKeever Kathleen , Kenrick Margaret , Leach Will , Polson Andrew , Saad Ola , Stephan Jean-Philippe , Shen Ben Quan , Tibbitts Jay , Yu Shang-Fan , Prabhu Saileta 
Published: April 2020

Abstract
CD22 is a B cell specific antigen and is expressed in Non-Hodgkin lymphoma (NHL) with limited expression on normal tissues. Currently, we are evaluating anti-CD22 antibody drug conjugates (anti-CD22 ADC) consisting of a humanized monoclonal IgG antibody conjugated to mitotic inhibitors such as DM1 (a maytansinoid), Monomethylauristatin F (MMAF), or Monomethylauristatin E (MMAE). The antibody binds to human and cynomolgus monkey CD22, but does not bind to rodent CD22. The cytotoxic drugs were conjugated to the antibody via uncleavable linkers (MCC-DM1, MC-MMAF) or cleavable linkers (SPP-DM1, vc-MMAE). These conjugates have demonstrated anti-tumor activity in mouse xenograft models of human B-cell tumors. This study was to assess and to compare the antigen-independent pharmacokinetics of anti-CD22 ADCs with uncleavable linkers (anti-CD22-MCC-DM1 and anti-CD22-MC-MMAF) and cleavable linkers (anti-CD22-SPP-DM1 and anti-CD22-vc-MMAE) in rats, and to support a pilot rat safety study. Following a single IV dose, the anti-CD22 ADCs exhibited a biphasic disposition profile. The conjugates with cleavable linkers had faster clearance compared to those with uncleavable linkers (59.4 mL/day/kg for anti-CD22-SPP-DM1 vs. 18.0 mL/day/kg for anti-CD22-MCC-DM1). The faster clearance for anti-CD22 ADCs with cleavable linker has also been observed with other ADCs (different target). In addition, it was found that free DM1 in plasma that was deconjugated from anti-CD22-SPP-DM1 had ∼70 fold higher exposure compared with that from anti-CD22-MCC-DM1 (area under curve (AUC) for free DM1 in plasma: 562 day*ng/mL for anti-CD22-SPP-DM1 vs. 8.18 day*ng/mL for anti-CD22-MCC-DM1). Furthermore, anti-CD22-SPP-DM1 had a different DM1 release profile compared with anti-CD22-MCC-DM1 when incubated with human plasma in vitro. Maximum concentrations of free DM1 were reached at ∼6 hr and ∼24 hr for anti-CD22-SPP-DM1 and anti-CD22-MCC-DM1, respectively. These observations were consistent with the safety findings in rats: anti-CD22-SPP-DM1 had greater toxicity (body weight loss, renal toxicity) compared with anti-CD22-MCC-DM1. The data described here indicate that the anti-CD22 ADCs with uncleavable linkers are cleared slower than, and have a better safety profile than the conjugates with cleavable linkers.
De Novo Expression of ATP Binding Cassette C1 (ABCC1) on T-ALL Cells Confers Resistance to Vincristine Independent of an MDR1 Phenotype.
Authors: Winter Stuart S. , Khawaja Hadya M. , Lovato Debbie M. , Larson Richard S. 
Published: April 2020

Abstract
Acquired drug resistance eventually leads to treatment failure in T-cell acute lymphoblastic leukemia (T-ALL). To define mechanisms of vincristine (VCR)-resistance, and to address whether inducible resistance in T-ALL might be mechanistically-dependent on ABC drug transporter expression, we developed VCR-resistant Sup T1 and Jurkat by incrementally growing cells in increasing drug concentrations. Jurkat (CD1a−, sCD3+) and Sup T1 (CD1a+, sCD3−) are arrested at “mature” and “cortical” stages of T-cell development and, respectively, showed 1190-fold (0.0006 μM to 0.714 μM) and 790-fold (0.0006 μM to 0.474 μM) increased resistance to VCR. Microarray analysis showed that expression of the ATP Binding Cassette C1 (ABCC1; Multidrug Resistance Protein, MDR2) was increased more than 29-fold in VCR-resistant cells as compared to the parental controls; fold increases in mRNA of ABCC1 were confirmed by RT-PCR, and cell surface expression by flow cytometric analysis. We then employed a flow cytometry-based assay that measures the ability of ABC pumps to extrude fluorescent dyes (Calcein AM). We confirmed that VCR-resistant Jurkat and Sup T1 actively extrude Calcein AM; drug extrusion was also effectively blocked by verapamil and other known ABCB1 antagonists. In contrast to our previous observations for ABCB1-mediated multidrug resistance (Estes, et al., British Journal of Haematology, in press), ABCC1-mediated drug resistance conferred a more narrow spectrum of multidrug resistance for the chemotherapeutic agents commonly used in T-ALL induction. To investigate whether ABC expression might impact patient outcome, we next examined ABCC1 and ABCB1 expression (&gt;1.5-fold) in 80 T-ALL patients treated on COG studies 8704 and 9404. Different from our earlier findings that ABCB1 expression in diagnostic marrows conferred a 100% risk for on-therapy relapse, we found that ABCC1 expression only predicted treatment failure in approximately 25% of affected patients. These results suggest that ABC transporters are differentially regulated in Sup T1 and Jurkat, and that substrate-specific ABC protein regulation results in unique multidrug resistance phenotypes in T-ALL. This study contributes to the repertoire of substrate-specific, drug resistant T-ALL cells lines that are available for modeling treatment failure, provides mechanistically-based insights into relapse, and establishes an experimental means for investigating compounds that reverse ABCC1 mediated drug resistance.
Multi-Drug Resistant Leukemic Cells Highly Express HLA Class I Molecules and Single-Chain Fv Diabody Specific to HLA-A Overcomes Drug Resistance in These Cells.
Authors: Jalili Ali , Ozaki Shuji , Kimura Naoki , Abe Masahiro , Matsumoto Toshio 
Published: April 2020

Abstract
Many tumor cells become resistant to commonly used cytotoxic drugs due to the overexpression of ATP-binding cassette (ABC) transporters. Specifically, p-glycoprotein (MDR-1) is frequently up-regulated in chemotherapy-resistant tumor cells, which is associated with poor prognosis. On the other hand, human leukocyte antigen (HLA) class I molecules are known to be significantly down-regulated in advanced tumor cells to escape from immune surveillance. However, the relationship between MDR-1 expression and HLA expression is not fully understood. Recently, we have developed a recombinant single-chain Fv diabody specific to HLA-A and demonstrated that this agent mediates cell death in HLA-overexpressing lymphoid tumor cells but not in normal cells (Cancer Res2007; 67:1184). Here, we investigated the expression levels of HLA class I in chemo-resistant leukemic cells and evaluated the therapeutic potential of single-chain Fv diabody specific to HLA-A, C3B3-DB (Chugai Pharmaceutical Co. Ltd., Tokyo, Japan). Chemotherapy-resistant cells were established by subculturing of myeloid leukemia cell line HL60 and Burkitt’s lymphoma cell line BL-TH in increasing doses of vincristine (VCR), and named HL60/VCR and BL-TH/VCR, respectively. MDR-1 is strongly expressed in HL60/VCR and BL-TH/VCR cells both at the mRNA and protein levels, but not in the parental cells. Interestingly, expression levels of HLA class I molecules are 8 times higher in HL60/VCR and BL-TH/VCR cells than in the parental cells, suggesting that MDR-1 modulates cell surface expression of HLA by its transporter function. Next, we examined the cytotoxic activity of C3B3-DB on these chemo-resistant cell lines. C3B3-DB induced apoptosis in HL60/VCR and BL-TH/VCR cells and these chemo-resistant cell lines were more sensitive to C3B3-DB than the parent cells. Combination of C3B3-DB with chemotherapeutic agents such as VCR and daunorubicin (DNR) resulted in enhanced cytotoxicity against HL60/VCR and BL-TH/VCR cells. Importantly, pretreatment of these chemo-resistant cell lines with C3B3-DB reduced expression levels of MDR-1 and increased drug retention in these cells as detected by flow cytometry and confocal microscopy. Furthermore, combination of C3B3-DB with VCR significantly blocked the cell cycle at the G2 phase compared with VCR alone. Similar results were obtained with primary acute myeloid leukemia cells from 2 patients, resulting in up-regulation of both HLA class I and MDR-1 molecules at relapse phase compared at diagnosis. These results suggest that C3B3-DB enhances cytotoxicity of chemotherapeutic agents and provides a novel approach for overcoming drug resistance in hematological malignancies.
c-Jun N-Terminal Kinase (JNK) Activation Failure Confers a New Mechanism of Anthracycline Resistance in Acute Myeloid Leukemia (AML).
Authors: Lagadinou Eleni D. , Ziros Panagiotis G. , Tsopra Olga A. , Dimas Kostas , Pantazis Panagiotis , Spyridonidis Alexandros , Zoumbos Nicholas C. 
Published: April 2020

Abstract
Chemotherapy resistance remains a major challenge in the treatment of acute myeloid leukemia (AML). Besides the P-glycoprotein efflux of chemotherapeutics, additional cellular factors may contribute to drug resistance in AML. c- Jun N-terminal Kinase (JNK) is a protein kinase activated after exposure of cells to chemotherapeutic agents. Recently, studies in solid tumours have associated chemoresistance with failure of cancer cells to activate JNK. We asked whether drug resistance in AML is also attributed to intrinsic failure of the AML blasts to activate JNK. In vitro treatment of U937 AML cell line with anthracyclines induced a rapid and robust JNK phosphorylation and apoptosis. In contrast, the anthracyline-resistant derivative U937R cells showed no JNK activation after exposure to anthracyclines, also at doses that resulted in high accumulation of the drug within the cells. Inhibition of JNK in drug-sensitive U937 cells made them resistant to anthracyclines. First, JNK1-siRNA transfected U937 cells exhibited a 50.4% and 61.3% reduced daunorubicin- (DNR, 1μM 24hr) and doxorubicin- (DOX, 1.5 μM 24hr) induced apoptosis respectively; as compared to empty vector or untransfected U937 cells (P&lt; 0.001). Second, pretreatment of U937 cells with the 420116 cell-permeable JNK inhibitor (1 μM) reduced to a less but yet significant extent DNR-induced apoptosis as compared to cells treated with a negative control peptide (P = 0.013&lt;0.05). On the other hand, selective restoration of the inactive JNK pathway in resistant U937R cells by transfection with a mutant form of the SEK1/MKK4 upstream activator of JNK sensitized U937R cells to anthracyclines, compared to empty vector transfected cells (3.3-fold increase in DNR-induced apoptosis, 1μM DNR, 24hr and 3.1-fold increase in DOX-induced apoptosis, 1.5μM DOX, 24hr). Furthermore, we assessed the activation of JNK pathway in 29 primary AML bone marrow samples after short term (30-60 min) in vitro exposure to DNR (1 μM) and correlated it with clinical data. We found a strong correlation between the in vitro pharmacodymanic changes of phospho-JNK levels in AML primary blasts and the response of the AML patients to standard induction chemotherapy (P = 0.012). In addition, the drug-induced JNK activation pattern correlated with AML evolving from antecedent MDS (P = 0.017) and patient age (P = 0.046). In summary, our in vitro and in vivo results suggest that JNK activation failure is another mechanism of anthracycline resistance in AML. Elucidating the ultimate mechanisms leading to JNK suppression in chemoresistant AML may be of major therapeutic value.
The Coexpression of AML1-ETO9a Isoform and AML1-ETO in Acute Myeloid Leukemia M2 Subtype and Its Clinical Significance.
Authors: Zhang Hong , Chen Zi-xing , He Jun , Miao Yu-qin , Cen Jian-nong , Qiu Qiao-chen 
Published: April 2020

Abstract
Previous studies on either animal model or clinical settings have demonstrated that the expression of AML1/ETO9a (AE9a), a truncated version of AML1-ETO (AE) generated by alternative splicing during transcription, in patients with acute myeloid leukemia M2 subtype may have clinical significance. Moreover, the simultaneously expression of both AE and AE9a may even be more meaningful to the prognosis of the patients. To validate this hypothesis, we have measured the mRNA level of both AE and AE9a in each individual M2 patient by quantitative real-time RT-PCR, and correlated the ratio of AE/AE9a mRNA levels with the clinical outcome and prognosis in these patients. In 35 newly-diagnosed AML-M2 patients with t(8;21), expression of AE9a was detected in 30 cases (85.71%). The median value of the percentages of AE9a and AE within overall transcripts of the fusion gene in these patients were 18.16% and 81.84%, respectively (p&lt;0.05). A positive correlation between the mRNA level of AE9a and AE was shown in 45 bone marrow samples from 19 patients during the follow-up, and the correlation coefficient was 0.900(P&lt;0.01). The expression level of both AE9a and AE decreased in 16 patients after one course of standard conventional chemotherapy. Among them, a reduced percentage of AE9a mRNA level was prominent in 11 (68.75%) patients, while the percentage of mRNA level of AE decreased in 5 (31.25%) cases. The complete remission rate after first course of chemotherapy was 81.82%(9/11)in 11 patients, and 20.00%(1/5)in 5 patients, suggesting a higher CR rate in those patients who exhibited more conspicuous decrease in mRNA level of AE9a (P&lt;0.05). The percentage of AE9a mRNA in the total fusion transcripts rose in all of the 5 relapsing cases. In a retrospective study on 92 patients of AML-M2 including 50 cases with t(8;21) and 42 case with either normal karyotype or other chromosome abnormalities, expression of AE9a has been found in these 50 cases by conventional RT-PCR. Between the AE9a positive and negative groups, no statistically significant difference had been found in complete remission rate, relapsing rate and the first CR period over a follow-up with medium time of 13 months (4–76 mths). However, 22 months of medium survival in AE9a positive group (38 cases) was remarkably shorter than 70 months in AE9a negative group (25 cases). Our results indicated that the mRNA of AE9a at a relatively lower level co-expressed with AE in most AML-M2 patients with t(8;21) translocation. The expression level of AE9a rather than AE may play more crucial role in development and progress of leukemia. AE9a positive patients have distinctly shorter survival time than the negative cases, predicting a worse prognosis.
Novel C-KIT Transcripts Identified in Mast Cell Leukemia: An Update of the Full Transcript and It’s Distribution.
Authors: Ozer Ozden , Zhao Yi D. , Ostler Kelly R. , Akin Cem , Anastasi John , Vardiman James , Godley Lucy 
Published: April 2020

Abstract
Introduction: Mast cell leukemia (MCL) is the rarest but most fatal form of mast cell neoplasms. Several C-KIT activating mutations have been identified in systemic mastocytosis (SM), the most frequent one being D816V. Little is known about C-KIT mutations in MCL. In 2005, at ASH we presented a novel C-KIT transcript in two patients with MCL, accounting for the sole C-KIT transcript in one of the patients. The novel form had a deletion of exons 12 and 13, encoding the N-terminal portion of the bipartite kinase domain. Here, we describe the complete C-KIT transcript, its response to stem cell factor (SCF) stimulation in vitro, and its expression in non- MCL samples.
Materials&Methods: The entire C-KIT coding region was amplified by RT-PCR from cDNA prepared from bone marrow (BM) samples of 2 patients with MCL, 2 patients with SM associated with acute myeloid leukemia, isolated CD34+ cells from healthy individuals, and 2 sets of purified mast cells from patients with SM. Several clones from each sample were fully sequenced. Its expression in patient samples was verified by Western blotting. An expression construct was made, and the mutant C-KIT was expressed in COS-1 cells. Western blotting was used to test the ability of the mutant protein to undergo autophosphorylation upon SCF stimulation.
Results: Complete sequencing demonstrated that the mutant C-KIT contained several mutations: a large deletion of amino acids 59–437, corresponding to loss of the first three immunoglobulin-like domains in the extracellular region, deletions of amino acids 510–513, deletions of the amino acids encoded by exons 12 and 13, and novel single amino acid deletions unique to each patient. The mutant C-KIT transcript was present in normal CD34+ cells and in 2 BMs with SM associated with AML. However, it was absent in purified mast cells from 2 patients with SM. In contrast, these displayed the widely recognised D816V point mutation. Western blotting from patient samples revealed a truncated protein of the predicted 59KD size. However, the novel C-KIT did not autophosphorylate upon stem cell factor stimulation.
Discussion: We have identified a novel C-KIT isoform. It was exclusively present in MCL but completely absent in purified mast cells of SM. These findings indicate that it is a marker of immaturity and aggressiveness among mast cell neoplasms, which may serve as a diagnostic and prognostic parameter. Its presence in non-neoplastic samples in similar amounts with the wild type suggest that it is not malignancy specific. However, its predominace over the wild type may correlate with a pathologic state that remains to be explored in other hematopoetic neoplasms.
The Inflammatory Subset of Monocytes Stimulates Angiogenesis at Sites of Ischemia by Altering the Balance of Pro- and Anti-Inflammatory Cytokines.
Authors: Gregory Alyssa D. , Capoccia Benjamin J. , Link Daniel C. 
Published: April 2020

Abstract
Our laboratory has previously shown that early delivery of monocytes to ischemic tissue by mobilization from the bone marrow to blood or by adoptive transfer enhances revascularization after acute vascular injury. There is accumulating evidence that monocytes represent a heterogeneous population, containing subsets with distinct functional properties. Herein, we show that the inflammatory subset of monocytes mediates the angiogenic response in a murine model of hindlimb ischemia induced by femoral artery excision (FAE). Inflammatory (CX3CR1loGr-1+) and resident (CX3CR1hiGr-1−) monocytes were sorted from the bone marrow of CX3CR1+/GFP mice and injected intravenously into recipient mice 24 after FAE. Revascularization was monitored by laser Doppler imaging. A marked improvement in blood flow was observed after adoptive transfer of inflammatory but not resident monocytes [ratio of blood flow in ischemic to nonischemic limb 7 days after surgery: 0.75±0.05 (inflammatory monocytes); 0.40±0.07 (resident monocytes); 0.40±0.03 (saline control); P &lt;0.01]. Consistent with this finding, inflammatory monocytes are selectively recruited to sites of ischemia. Finally, we show that the bone marrow is a rich reservoir of inflammatory monocytes that is readily mobilized into the blood by ischemia. To begin to elucidate the mechanisms by which inflammatory monocytes stimulate angiogenesis, we determined the effect of the adoptive transfer of monocytes on the local production of key cytokines/chemokines in the ischemic tissue. Specifically, a non-biased multiplex immunoassay-based screen for soluble factors present in the cell-free muscle homogenates was performed. Data revealed decreased levels of the pro-inflammatory cytokines IL-6, KC, and mip-1beta and increased levels of the anti-inflammatory cytokine IL-10 in mice receiving an adoptive transfer of monocytes. In keeping with this observation, levels of free myoglobin were dramatically decreased in mice receiving an adoptive transfer of monocytes, indicating a reduction in inflammation-mediated tissue damage [myoglobin levels at 28 hours post-ischemia: 18,580±879 ng/mL (no adoptive transfer); 158±112 ng/mL (adoptive transfer)]. Interestingly, known regulators of angiogenesis, MCP-1, MMP-9, VEGF, and FGF-basic were also significantly increased. Taken together, these data indicate that adoptive transfer of a small number of monocytes shifts the balance of pro- and anti-inflammatory signals in the ischemic environment, which limits tissue damage and enhances revascularization. This finding has important therapeutic implications, since deliver of inflammatory monocytes to sites of ischemia may polarize the microenviroment towards tissue regeneration and angiogenesis.