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  • American Society of Hematology  (15)
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  • American Society of Hematology  (15)
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  • 1
    In: Blood, American Society of Hematology, Vol. 132, No. Supplement 1 ( 2018-11-29), p. 2960-2960
    Abstract: Introduction: Diffuse large B-cell lymphoma (DLBCL) and follicular lymphoma (FL) are the most common types of B-cell non-Hodgkin's lymphoma (B-NHL). R-CHOP (rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisolone) is the frontline treatment for primary DLBCL and FL. Pegylated liposomal doxorubicin (PLD) is the liposome formulation of doxorubicin. R-CDOP, in which PLD was used to substitute for doxorubicin, has been recommended for patients with poor cardiac function. This study was aimed to elucidate the therapy responses and cardiotoxicity with R-CDOP and R-CHOP observed in frontline treatment of patients with primary DLBCL and FL. Methods: From January 2015 to January 2018, 77 newly diagnosed DLBCL and 25 newly diagnosed FL patients were treated with R-CDOP or R-CHOP regimen randomly. Among the 102 patients, 67 cases were treated with R-CDOP and 35 cases received R-CHOP therapy. The regimen of R-CDOP or R-CHOP consisted of Rituximab 375 mg/m2 on cycle day 0, and along with Cyclophosphamide 750 mg/m2, PLD 35 mg/ m2 or Epirubicin 70 mg/m2 (substituting for Doxorubicin), Vindesine 3.0 mg/m2 on cycle day 1 and Prednisone 100mg/d from day1 to day5 of each cycle. This regimen was implemented every 21 days approximately. After 3 cycles, tumor response were assessed and classified as complete response (CR), partial response (PR), stable disease (SD), or progressive disease (PD) according to the Cheson criteria. Cardiac function was also evaluated, including N-terminal pro B-type natriuretic peptide (NT-proBNP), left ventricular ejection fractions (LVEF) and myocardial enzymes. Patients with a therapeutic assessment of CR/PR received another 3 cycles of R-CDOP or R-CHOP regimen. Patients whose responses were evaluated for SD/PD were withdrawn from our study. After 6 cycles, tumor responses and cardiac functions were assessed again. Then the patients were followed up every three months up to 9 months or till PD. The analyses of data were performed using SAS (statistical analysis system) 9.3. Repeated measurement of variance analysis was used to determine the statistical significances of rate differences between R-CDOP group and R-CHOP group. Other statistical data analyses were performed using the two-tailed Student's t test. All differences were considered to be statistically significant when the P value was less than 0.05. Results: The clinical characteristics were summarized in Table 1. All the patients were randomized to one of the two treatment arms. Baseline characteristics had no significant difference between the two groups. The follow up time was range from 3 to 9 months. In R-CDOP group, the CR/PR ratio was higher than that in R-CHOP group (P 〈 0.05, Table 2). For DLBCL patients, CR/PR ratio in R-CDOP group was also higher that in R-CHOP group (P 〈 0.05, Table 3.1). However, for FL patients, only CR ratio in R-CDOP group was higher than that in R-CHOP group (P 〈 0.05, Table 3.1). For young patients (age≤60 years) and patients whose stage was I/II, CR/PR ratio in R-CDOP group was also higher than that in R-CHOP group (P 〈 0.05, Table 3.2 and 3.3). However, for old patients (age 〉 60years) and patients whose stage was III/IV, the CR/PR ratio in R-CDOP group and R-CHOP group showed no statistical difference (Table 3.2 and 3.3). NT-proBNP, LVEF and myocardial enzymes were measured to evaluate cardiac function. Our study showed that NT-proBNP levels had significant difference after 3 and 6 cycles treatment between R-CDOP group and R-CHOP group (62.7±44.8 pg/ml, n=29 vs. 101.0±65.0 pg/ml, n=26 and 55.6±35.6 pg/ml, n=25 vs. 103.20±57.7 pg/ml, n=25, P 〈 0.05). Surprisingly, the difference between these two groups disappeared in follow-up period. LVEF (0.66±0.02, n=18 vs. 0.64±0.01, n=12) and myocardial enzymes, including creatine kinase (CK, 65.1±25.1 U/L, n=27 vs. 127.6±64.1 U/L, n=20) and creatine kinase isoenzyme CKMB (8.0±2.5 ng/ml, n=24 vs. 14.9±9.3 ng/ml, n=17), were found to have significant differences in R-CDOP group and R-CHOP group at 9 months after the last cycle (P 〈 0.05). Conclusions: Our findings in this study indicated that the R-CDOP regimen offers superior oncologic efficiency when compared to standard R-CHOP regimen in primary DLBCL and FL patients. This study also showed that the substitution of conventional doxorubicin by PLD was a safer therapeutic option for patients' cardiac function. Disclosures No relevant conflicts of interest to declare.
    Type of Medium: Online Resource
    ISSN: 0006-4971 , 1528-0020
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    RVK:
    Language: English
    Publisher: American Society of Hematology
    Publication Date: 2018
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  • 2
    In: Blood, American Society of Hematology, Vol. 138, No. Supplement 1 ( 2021-11-05), p. 4513-4513
    Abstract: Introduction: Waldenström macroglobulinaemia/ Lymphoplasmacytoid lymphoma (WM/LPL) is a rare lymphoproliferative neoplasm characterized by small B lymphocytes proliferation. Abnormalities of thyroid hormones are common in clinical courses. Yet, the role of thyroid complications has not been explored in WM/LPL. Hence, the aim of this study was to investigate the clinical significance of thyroid complications in WM/LPL. Methods: 105 clinically diagnostic WM/LPL patients from Shandong Provincial Hospital were enrolled with informed consents. Baseline and clinical data concerning sex, age, International Staging System Waldenstrom Macroglobulinemia (ISSWM) score et al were collected. Chi-square test was used for comparison of clinical characteristics. The Kaplan-Meier method was used for analysis of survival outcomes. Cox regression analyses were utilized to identify prognostic-related key factors associated with overall survival (OS) and progression-free survival (PFS) in WM/LPL patients. Microarray datasets GSE6691 were obtained from Gene Expression Omnibus. Results: Over the 105 WM/LPL patients, the median overall survival (OS) was not reached and median progression-free survival (PFS) was 96 months (Figure 1A, 1B). Patients classified as complete response (CR)/ partial response (PR)/ stable disease (SD), showed better OS and PFS than patients with progression disease (PD) (Figure 1C, D). There were 13.3% of enrolled patients with mixed thyroid complications. The results of Chi-square test showed that thyroid complications were significantly associated with reduced IgM level (p=0.036) and elevated β2-macroglobulin (p=0.032). Moreover, patients without thyroid comorbidities were more likely to get overall response (CR+PR) to the first-line treatment (p=0.004). Kaplan-Meier curves showed patients with thyroid complications had significantly shorter OS (p=0.02) and PFS (p & lt;0.001) versus those without thyroid complications (Figure 1E, F). In the univariate Cox regression model, age (p=0.022), ISSWM score (p=0.014) and thyroid complications (p & lt;0.001) were significantly associated with an increased risk of progression developed. Subsequent multivariate analysis showed the presence of thyroid complications was an independent prognostic indicator for PFS in WM/LPL patients (p=0.03). However, there was no statistical significance of thyroid complications in OS. Microarray dataset analysis was conducted to further investigate the role of thyroid-related genes in WM/LPL patients. A network of interactions among thyroid-related genes and critical factors in WM/LPL, including MYD88 and CXCR4, was shown in Figure 1G. Correlations were statistically significant between SLC5A5 (p & lt;0.05), TG (p & lt;0.01), TPO (p & lt;0.01) and CXCR4 by Spearman correlation analysis (Figure 1H, I). In addition, differential gene expression analysis between the WM and normal lymphocytes was assessed (Figure 1J). Thyroid-related genes with statistical significance were annotated in the volcano plots (Figure 1K). Enrichment analysis indicated that differential genes were involving in PI3K-Akt signaling pathway and response to peptide hormone (Figure 1L). Moreover, five of them reached statistical significance, illuminating the potential importance of thyroid-related genes in WM/LPL (Figure 1M). Conclusion: Taken together, the present study was the first investigation on the role of thyroid complications in WM/LPL. Patients with thyroid complications showed worse clinical characteristics and conferred independent prognostic significance. The primary strength of this study is that it provides robust real-world evidence on the prognostic role of thyroid complications, highlighting the need to monitor and appropriately manage WM/LPL patients with thyroid complications in medical admissions. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.
    Type of Medium: Online Resource
    ISSN: 0006-4971 , 1528-0020
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    RVK:
    Language: English
    Publisher: American Society of Hematology
    Publication Date: 2021
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  • 3
    In: Blood, American Society of Hematology, Vol. 132, No. Supplement 1 ( 2018-11-29), p. 5781-5781
    Abstract: Objective: Retrospectively analysing the outcome of additional medium-dose etoposide (VP-16 30mg/kg) in allogeneic hematopoietic stem cell transplantation (allo-HSCT) for acute lymphoblastic leukemia (ALL). Methods: From 2010.10.1 to 2018.6.30, 53 ALL patients received allo-HSCT in Shandong provincial hospital. 20 conditioning with medium-dose VP-16 (15 in CR , 5 in refractory and/or relapse disease) (VP-16 group), 33 conditioning without medium-dose VP-16 (32 in CR and 1 in refractory and/or relapse disease) (control group) . The basic conditions of the primary disease, treatment-related toxicity, leukemia-free survival (LFS), overall survival (OS), graft-versus-host disease (GVHD) and relapse-free survival (GRFS), non-relapse mortality (NRM), relapse incidence (RI), GVHD and so on were compared between the two groups. Results: The VP-16 group included more R/R(refractory and/or relapse disease) patients and was associated with improved 3-year-LFS (70.7% vs. 56.5%, p= 0.565) and 3-year-OS (68% vs. 59.4%, p=0.825), although there was no statistical difference. Moreover, the additional of VP-16 lead to reduced incidence of aGVHD (II,III,IV) (10% vs. 30.7%, p=0.146) and cGVHD (9.1% vs 24%, p=0.25), resulting in improved GRFS (63.6% vs. 48.7%, P=0.343). RI was similar between the two group (29.3% vs 27.2%, p=0.76). Meanwhile, the addition of VP-16 enhanced the intensity of conditioning regimen, its NRM was relatively lower than control group (0% vs. 20.2%, P=0.371). Conclusion: The intensified conditioning regimen with medium-dose VP-16 did not increase NRM and GVHD, with tendency to increase OS and LFS, may become an effective pre-treatment scheme in allo-HSCT for ALL. Disclosures No relevant conflicts of interest to declare.
    Type of Medium: Online Resource
    ISSN: 0006-4971 , 1528-0020
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    RVK:
    Language: English
    Publisher: American Society of Hematology
    Publication Date: 2018
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  • 4
    In: Blood, American Society of Hematology, Vol. 136, No. Supplement 1 ( 2020-11-5), p. 17-18
    Abstract: Introduction N6-methyladenosine (m6A) is the most prevalent post-transcriptional modification of eukaryotic mRNA. Accumulating evidence suggests that RNA m6A methylation exerts crucial roles in oncogenesis. However, the mRNA m6A methylation pattern in chronic lymphocytic leukemia (CLL) has not been investigated. Hence, the aim of this study is to perform a comprehensive profiling to identify distinct m6A methylation signatures in CLL patients. Methods Peripheral blood samples from de novo CLL patients were collected with informed consents at the Department of Hematology in Shandong Provincial Hospital Affiliated to Shandong First Medical University. CD19+ B cells were isolated with informed consents from healthy donors. Methylated RNA immunoprecipitation sequencing (MeRIP-Seq) was conduct to profile mRNA m6A methylation of CLL-B cells and normal CD19+ B cells at Novogene (Beijing, China). The library preparations were sequenced on an Illumina Novaseq platform with a paired-end read length of 150 bp according to the standard protocols. The sequencing was carried out with 3 independent biological replicates. After mapping reads to the reference genome, exomePeak R package was used for the m6A peak identification in each anti-m6A immunoprecipitation group with the corresponding input samples serving as a control, and q-value threshold of enrichment of 0.05 was used for all data sets. The m6A-enriched motifs of each group were identified by HOMER. Differential peak calling was performed using exomePeak R package with parameters of p-value & lt; 0.05 and fold_change & gt; 1. Functional enrichment analyses of gene ontology (GO) and kyoto encyclopedia of genes and genomes (KEGG) of differentiated peaks associated genes were performed. All investigators comply with the guiding principles for experimental procedures found in the Declaration of Helsinki of the World Medical Association. Results By MeRIP sequencing, significant distribution of methylation peaks were detected in 5'UTR, 3'UTR and CDS regions of CLL primary cells (Figure 1A) and normal B cells (Figure 1B). Figure 1C-D illustrated the percentage of methylation peaks in the five regions, suggesting distinct m6A patterns in CLL cells. Moreover, Figure 1E-F revealed the positions of m6A methylation peaks in chromosomes of CLL and normal B cells. Furthermore, the compared distributions of m6A methylation peaks in CLL and normal B cells were presented in Figure 2A. Besides, the bean plot visibly displayed the obvious differentiation of methylation peaks in CLL group and normal B cells in read density (Figure 2B). Importantly, a total of 1836 significantly changed peaks, of which 1519 were significantly up-regulated and 317 peaks were significantly down-regulated (p & lt;0.05, |log2Foldchange| & gt;1; Figure 2C). These m6A peaks were located across 1850 genes. Functional enrichment analyses identified that differentiated peaks associated genes were potentially regulate RNA metabolic process via oncogenic pathways in CLL pathogenesis (Figure 3A-B). In addition, HOMER analysis identified 38 significant de novo m6A peak motifs, top 10 most significant peak motifs of which were presented (Figure 3C), illuminating potential detailed mechanism of m6A RNA methylation in the tumorigenesis and progression of CLL. Conclusion Taken together, our investigations explored for the first time the m6A methylation pattern of mRNA in CLL. m6A modifications play crucial roles in the progression and survival of CLL patients,highlighting m6A modifications-targeted intervention formulating a novel treatment paradigm in progressed CLL that warrants clinical investigation. Disclosures No relevant conflicts of interest to declare.
    Type of Medium: Online Resource
    ISSN: 0006-4971 , 1528-0020
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    RVK:
    Language: English
    Publisher: American Society of Hematology
    Publication Date: 2020
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  • 5
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    American Society of Hematology ; 2021
    In:  Blood Vol. 138, No. Supplement 1 ( 2021-11-05), p. 2857-2857
    In: Blood, American Society of Hematology, Vol. 138, No. Supplement 1 ( 2021-11-05), p. 2857-2857
    Abstract: Background : Eltrombopag is an agonist of the thrombopoietin receptor, and it has achieved great efficacy in the treatment of immunologic thrombocytopenic (ITP) and reduction of platelets (PLT) after hematopoietic stem cell transplantation (HSCT), but it remains unclear whether Eltrombopag can promote PLT engraftment after HSCT. Methods : 37 patients with HSCT were administered Eltrombopag 50 mg (qd) from +1 day until PLT & gt;50x10^9/L or 1 month after HSCT. 45 patients in the same period applied TPO to promote PLT engraftment after HSCT and served as a control group. The engraftment time of PLT and WBC (white blood cell) as well as the GVHD incidence and adverse reaction were compared and analyzed in the two groups. Results : A total of 37 patients received Eltrombopag, 2 of the patients withdrew from the study due to intolerable gastrointestinal reactions. 35 patients eventually completed treatment with Eltrombopag. In Eltrombopag group, median time to WBC engraftment was 12 days (range, 10-17 days) and PLT engraftment was 15 days (range, 10-29 days). At the 1st month after HSCT, the median WBC was 3.93 x10 9/L (range, 0.87-40.01 x10 9/L) and the median PLT was 91 x10 9/L (range, 33-401 x10 9/L) in the Eltrombopag group. MKCs in the bone marrow were assayed at 1 month after HSCT. Median MKCs level was 40 (range, 2-345) in the Eltrombopag group. 13 (37%) patients developed grade II-IV aGVHD and 1 (2%) patient developed cGVHD in the Eltrombopag group. Of these 35 patients, 3 (8%) presented with elevated total bilirubin (TBIL), no patient had elevated alkaline phosphatase (ALP), 17(48%)developed proteinuria, 1 (2%) people with elevated blood urea nitrogen (BUN) and no patient showed elevated creatinine (CREA). A total of 45 patients were applied TPO during the same period. Among these patients, the median engraftment time for WBC and PLT was 12 days (range, 9-23 days) and 15 days (range, 9-41 days), respectively. At the 1st month after HSCT, the median number of WBC and PLT were 4.30 x10 9/L (range, 0.99-23.63 x10 9/L) and 79.5 x10 9/L (range, 5-512 x10 9/L), respectively. At the 1st month after transplantation, the median number of MKCs in the bone marrow were 44 (range, 2-788). 16 (35%) patients developed grade II-IV aGVHD and 6 (13%) patients developed cGVHD in the TPO group. Among these 45 patients, 7 (15%) presented with elevated TBIL, 5 (11%) had elevated ALP, 25 (55%) developed proteinuria, 4 (8%) people with elevated BUN and 1 (2%) patient showed elevated CREA. The engraftment time of PLT and WBC were same in the Eltrombopag group and the TPO group and the adverse reactions occurred similarly in both groups. Conclusion : Eltrombopag can be used to promote PLT engraftment after all-HSCT with acceptable adverse reaction. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.
    Type of Medium: Online Resource
    ISSN: 0006-4971 , 1528-0020
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    RVK:
    Language: English
    Publisher: American Society of Hematology
    Publication Date: 2021
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  • 6
    In: Blood, American Society of Hematology, Vol. 134, No. Supplement_1 ( 2019-11-13), p. 5323-5323
    Abstract: Introduction: B-cell non-Hodgkin lymphoma is the most frequent type of non-Hodgkin's lymphoma. RCHOP regimen is established as the standard therapy for aggressive and indolent B-cell NHL, which has a 10%-20% rate of febrile neutropenia (FN). Recently, pegylated recombinant human granulocyte colony stimulating factor (PEG-rhG-CSF) is frequently used in clinical practice. This randomized controlled clinical study was conducted to investigate the efficacy and safety of prophylactic PEG-rhG-CSF in patients with B-cell non-Hodgkin lymphoma on RCHOP chemotherapy. Methods:We included 162 patients with pathological diagnosis of B-cell non-Hodgkin lymphoma including diffuse large B-cell lymphoma, follicular lymphoma and mantle cell lymphoma (MCL),from October 2016 to May 2019 at Shandong Provincial Hospital Affiliated to Shandong University. All patients gave written informed consent in accordance with the Declaration of Helsinki. The patients were randomized into PEG-rhG-CSF and rhG-CSF groups. Each patient received three cycles of chemotherapy with identical RCHOP regimens. In the study group, the patients received PEG-rhG-CSF 6mg(weight≥45Kg)or 3mg(weight≤45Kg)once 24 hours after the end of chemotherapy drugs of every chemotherapy cycle. In the control group, they weren't preventively administered rhG-CSF. If their neutrophil count (ANC)≤1.0×109/L, they were administered rhG-CSF:5ug/kg/day until their neutrophil count (ANC)≥2.0×109/L. The primary endpoint was the incidence of III/IV grade neutropenia and febrile neutropenia(FN) after each chemotherapy cycle. Meanwhile the rate of antibiotics application and safety were observed. Analyses were performed with SPSS Statistics 20.0 (IBM-SPSS, Chicago, Illinois). The numerical data was presented as mean ± SD. Statistical analysis was performed using one-way analysis of variance and chi-square test. A p-value 〈 0.05 was considered statistically significant. Results: Clinical characteristics for PEG-rhG-CSF and rhG-CSF groups were shown in Table1. There were no significant differences in age, gender,height, body weight, body mass index, Ann Arbor and IPI staging. The incidence of IV grade neutropenia during cycle 1 in 81 evaluable study cycles and 81 evaluable control cycles were 7.41% and 35.80%( P 〈 0.01), with durations of 2.85±0.62 days and 3.11±1.23 days (P 〉 0.05). The differences in I/II/III grade neutropenia between study and control groups weren't statistically significant (Table2,Fig.1). After secondary prophylactic use of PEG-rhG-CSF In the study group, the incidences of III/IV grade neutropenia decreased from 77.78% to 14.81% (P 〈 0.01).Statistically significant differences were observed in the incidences of FN (12.35% and 34.57% for the PEG-rhG-CSF and rhG-CSF groups, respectively; P 〈 0.01) and in the proportion of patients who received antibiotic therapy (11.11% and 37.04%, respectively; P 〈 0.01) during cycle 1(Table2,Fig .2). The safety profiles of PEG-rhG-CSF and rhG-CSF were similar. Bone pain occurred in 7.41% of the cases during the study cycles and 2.47% in the control cycles (P 〉 0.05 ), which were mostly mild or moderate. Patients receiving PEG-rhG-CSF who developed III/IV grade neutropenia were significantly older than those without neutropenia (53.41±14.96 vs. 63.64±4.65;years; p=0.01) (Fig.3).The incidence of III/IV grade neutropenia in patients older than 60 years was significantly higher than that in patients younger than 60 years(24.44% vs. 6.38%; P =0.038). Conclusions: Prophylactic use of PEG-rhG-CSF could effectively reduce the incidences of grade III/IV neutropenia and FN, which ensures that patients with lymphoma receive standard-dose chemotherapy to improve prognosis. III/IV grade neutropenia after prophylactic use of PEG-rhG-CSF were more likely to occur in patients older than 60 years. After the use of PEG-rhG-CSF, the elderly patients should be pay more attention to them. Disclosures No relevant conflicts of interest to declare.
    Type of Medium: Online Resource
    ISSN: 0006-4971 , 1528-0020
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    Language: English
    Publisher: American Society of Hematology
    Publication Date: 2019
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  • 7
    In: Blood, American Society of Hematology, Vol. 116, No. 26 ( 2010-12-23), p. 6114-6122
    Abstract: Ectopically expressed, human B-domainless (hB) factor 8 (F8) in platelets improves hemostasis in hemophilia A mice in several injury models. However, in both a cuticular bleeding model and a cremaster laser arteriole/venule injury model, there were limitations to platelet-derived (p) hBF8 efficacy, including increased clot embolization. We now address whether variants of F8 with enhanced activity, inactivation resistant F8 (IR8) and canine (c) BF8, would improve clotting efficacy. In both transgenic and lentiviral murine model approaches, pIR8 expressed at comparable levels to phBF8, but pcBF8 expressed at only approximately 30%. Both variants were more effective than hBF8 in cuticular bleeding and FeCl3 carotid artery models. However, in the cremaster injury model, only pcBF8 was more effective, markedly decreasing clot embolization. Because inhibitors of F8 are stored in platelet granules and IR8 is not protected by binding to von Willebrand factor, we also tested whether pIR8 was effective in the face of inhibitors and found that pIR8 is protected from the inhibitors. In summary, pF8 variants with high specific activity are more effective in controlling bleeding, but this improved efficacy was inconsistent between bleeding models, perhaps reflecting the underlying mechanism(s) for the increased specific activity of the studied F8 variants.
    Type of Medium: Online Resource
    ISSN: 0006-4971 , 1528-0020
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    Language: English
    Publisher: American Society of Hematology
    Publication Date: 2010
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  • 8
    In: Blood, American Society of Hematology, Vol. 120, No. 24 ( 2012-12-06), p. 4829-4839
    Abstract: Bcr-Abl tyrosine kinase inhibitors (TKIs) have been a remarkable success for the treatment of Ph+ chronic myeloid leukemia (CML). However, a significant proportion of patients treated with TKIs develop resistance because of leukemia stem cells (LSCs) and T315I mutant Bcr-Abl. Here we describe the unknown activity of the natural product berbamine that efficiently eradicates LSCs and T315I mutant Bcr-Abl clones. Unexpectedly, we identify CaMKII γ as a specific and critical target of berbamine for its antileukemia activity. Berbamine specifically binds to the ATP-binding pocket of CaMKII γ, inhibits its phosphorylation and triggers apoptosis of leukemia cells. More importantly, CaMKII γ is highly activated in LSCs but not in normal hematopoietic stem cells and coactivates LSC-related β-catenin and Stat3 signaling networks. The identification of CaMKII γ as a specific target of berbamine and as a critical molecular switch regulating multiple LSC-related signaling pathways can explain the unique antileukemia activity of berbamine. These findings also suggest that berbamine may be the first ATP-competitive inhibitor of CaMKII γ, and potentially, can serve as a new type of molecular targeted agent through inhibition of the CaMKII γ activity for treatment of leukemia.
    Type of Medium: Online Resource
    ISSN: 0006-4971 , 1528-0020
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    Language: English
    Publisher: American Society of Hematology
    Publication Date: 2012
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  • 9
    In: Blood, American Society of Hematology, Vol. 136, No. Supplement 1 ( 2020-11-5), p. 2-3
    Abstract: Introduction: CD5-positive diffuse large B-cell lymphoma (CD5+ DLBCL) is characterized by a poor prognosis, poorly respond to the regulatory treatment strategy, and a relatively high incidence of central nervous system (CNS) infiltration. In this study, we aim to identify key differentially expressed miRNAs (DE-miRNAs) and their target genes in the peripheral blood of CD5+ refractory and relapsed (CD5+ R/R DLBCL) patients. The relationship of the DE-miRNAs and the pathogenesis of CD5+ R/R DLBCL will also be analyzed by bioinformatics tools. Methods: Three female patients with confirmed CD5+ R/R DLBCL were enrolled in this study, their age were 38, 62 and 65 years old, respectively. Three healthy female adults aged 42, 55 and 61, respectively were selected as the control group. The peripheral venous blood of them was collected for RNA extraction and standard small RNA sequencing. Differentially expressed miRNAs analysis was performed with R package edgeR. The target genes of DE-miRNAs were predicted by miRNet. A protein protein interaction (PPI) network was established for these target genes through string database. Functional annotation and pathway enrichment analyses for the target genes were performed through DAVID database to identify their potential functions, target genes, and pathways they might be involved in. Results: 1. Scatter plots, Volcano plots and Heat-maps were used to visualize miRNAs of Differentially expressed genes. As shown in Fig.1, Fig. 2 and Fig. 3. 2. Fifty-five sequences were significantly upregulated and 23 were significantly downregulated in patients with CD5+ R/R DLBCL.Among the candidate miRNAs, 11 up-regulated genes and 4 down-regulated genes were selected according to the log2FC value. The target genes of 11 potential up-regulated and 4 down-regulated DE-miRNAs were successively predicted by As shown in Table 1, a total of 439 and 632 predicted targets of the up-regulated and down-regulated DE-miRNAs were identified, respectively. 3. PPI networks of predicted target genes of 11 upregulated DE-miRNAs (Fig.4a) and 4 downregulated DE-miRNAs (Fig. 4b) were separately constructed using the STRING database and Cytoscape software. According to a degree, the top 10 hub genes in the networks were screened out and were listed inTable 2. Six important hub genes were identified, including two target genes predicted by up-regulating DE-miRNAs, namely NRAS and PIK3R1, and four target genes predicted by down-regulating DE-miRNAs, namely EGFR, VEGFA, IGF1 and Grb2. 4. DAVID now provides a comprehensive set of functional annotation tools for investigators to understand biological meaning. GO analysis was divided into three functional groups, including molecular function (MF), biological processes (BP), and cell composition (CC). The top 10 GO terms of targets of up-regulated DE-miRNAs were presented in Fig.5a-c. The top 10 GO terms of targets of down-regulated DE-miRNAs were shown in Fig. 5d-f. 5. Based on the KEGG database, we analyzed the pathways in which the differentially expressed target genes were involved in. As shown in Fig. 6a-b. The targets of up-regulated DE-miRNAs were enriched in pathways in cancer, oxytocin signaling pathway, ErbB signaling pathway, Rap1 signaling pathway, and proteoglycans in cancer. Whereas the targets of down-regulated DE-miRNAs were enriched in pathways in cancer, Ras signaling pathway, and PI3K-Akt signaling pathway. Conclusions: In this study, we analyzed the differentially expressed miRNAs in CD5+ R/R DLBCL patients, identified their potential functions, target genes, and pathways they might be involved in. This study found that ErbB signaling pathway, Rap1 signaling pathway, Ras signaling pathway and PI3K Akt signaling pathway were the most frequently involved pathways of miRNAs related genes. Target genes including NRAS, PIK3R1, EGFR, VEGFA, IGF1, and Grb2 might have a close relationship in the pathogenesis of CD5+ R/R DLBCL. New targeted drugs related to these pathways and genes may be beneficial to the treatment of CD5+ DLBCL. Our preliminary informatic results might be helpful to deeply understand the pathogenesis and chemotherapy resistance mechanism of CD5+ R/R DLBCL. In the future, we will verify our preliminary informatic results in pathological tissues from patients with CD5+ DLBCL in larger samples. Disclosures No relevant conflicts of interest to declare.
    Type of Medium: Online Resource
    ISSN: 0006-4971 , 1528-0020
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    Language: English
    Publisher: American Society of Hematology
    Publication Date: 2020
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  • 10
    In: Blood, American Society of Hematology, Vol. 132, No. Supplement 1 ( 2018-11-29), p. 179-179
    Abstract: Leukemias containing Mixed Lineage Leukemiagene rearrangement (MLL-r) account for 5-10% of human acute leukemia cases and are associated with a poor prognosis. The unmet clinical need and the lack of an effective targeted therapy emphasizes the need for novel approaches for these malignancies.Recent studies have discovered an essential role for the histone H3 lysine 79 (H3K79) methyltransferase DOT1L in the maintenance of MLL-r leukemias. Phase-I clinical trials (NCT01684150 and NCT02141828) have demonstrated the safety and clinical activity of the DOT1L-specific small molecule inhibitor EPZ5676 (Pinometostat, Epizyme Inc.). However, the variable response of the MLL-r patients in these clinical trials indicates the need to further understand the mechanism of action and resistance to DOT1L inhibitors. To address this issue, we designed a high-density CRISPR screen approach (Fig 1) to dissect the function of Dot1l coding regions in mouse MLL-AF9 leukemic cells, a well-established mouse model that mimics human MLL-r acute myeloid leukemia (AML). We constructed a library of 602 sgRNA that targets most of the "NGG" protospacer adjacent motifs (PAM) within Dot1l exons. The average targeting density is 7.7 bp per sgRNA (i.e. 2.5 a.a. per sgRNA). We delivered this sgRNA library into Cas9-expressing MLL-AF9 AML cells using lentiviral transduction, and then compared the frequencies of each integrated sgRNA sequence before vs. after 12 days of culture using high-throughput sequencing (NextSeq, Illumina Inc.). The results revealed a significant depletion of clusters of sgRNA targeting the known functional regions including the lysine methyltransferase (KMT) core and the AF9-binding domains of DOT1L. We mapped the CRISPR scan score to the previously solved three-dimensional structures of DOT1L and found the high-density CRISPR scan clearly distinguishes the substrate binding pocket from the less important regions within the KMT domain. Furthermore, we identified CRISPR hotspots that coincide with amino acid residues directly contacting the enzymatic substrate S-adenosylmethionine (SAM). Similarly, CRISPR scan identifies amino acid hot spots in DOT1L that establish a direct interaction with the cofactor AF9. These results implicate the utility of the high-density CRISPR scan to pinpoint the functional elements within a protein to a sub-domain resolution. To identify regions in DOT1L that mediate response of MLL-r leukemia to DOT1L-inhibitors, we conducted parallel CRISPR scans by culturing the Dot1l sgRNA library transduced MLL-AF9-Cas9+cells in either DMSO (vehicle) or the DOT1L inhibitor (1 uM EPZ5676) for 12 days. These paired screens revealed distinct patterns of the CRISPR scan scores between the two culture conditions. Surprisingly, a cluster of 31 sgRNA targeting the amino acid residues T560 - S661 of DOT1L was significantly enriched only in the presence of the DOT1L inhibitor.Expression of individual sgRNA targeting this region confirmed the EPZ5676-resistant phenotype observed in the CRISPR scans. Computational modeling of the structure of DOT1L suggests this newly identified region overlaps with two alpha-helixes motifs in a predicted coiled-coil domain. Finally, we examined eight clinically observed DOT1L missense mutations in this region (cBioPortal database; 54,510 patients) and foundthree DOT1L-mutant constructs (A591V, G594S and L626P) when expressed in the MLL-AF9 leukemia cells can confer resistance to EPZ5676. These findings suggest the utility of the high-density CRISPR scan for de novoidentification of drug-resistant mutant alleles in the human population, which maydirect future clinical decisions and benefit patients with specific genomic backgrounds. Disclosures No relevant conflicts of interest to declare.
    Type of Medium: Online Resource
    ISSN: 0006-4971 , 1528-0020
    RVK:
    RVK:
    Language: English
    Publisher: American Society of Hematology
    Publication Date: 2018
    detail.hit.zdb_id: 1468538-3
    detail.hit.zdb_id: 80069-7
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