bluuebunny/crossref_metadata_2025 huggingface dataset:
Tigecycline Was a Promising Candidate for Febrile Neutropenic and Carbapenem-Resistant Patients with Hematologic Malignancy
Authors: Xu Bing , He Lingli , Shi Pengcheng , Guo Xutao , Li Jie , Wang Yan , Deng Manman , Zhou Yong , Li Yin , Liang Jiabao
Published: October 2019

Background: To investigate the efficacy and safety of tigecycline (TGC) in febrile neutropenic and carbapenem-resistant patients with hematologic malignancy.

Methods: The clinical data of 71 patients with hematologic malignancy, febrile neutropenia (FN) were retrospectively analyzed. They are unresponsive to carbapenem for 3~5 days before receiving TGC. The loading dose of tigecyclin was 100 mg at the first time, and 50 mg every 12 hours as maintenance therapy. The efficacy was evaluated according to the Ministry of Health issued in 2007, meanwhile, the drug side effects were also evaluated.

Result: The median duration of neutropenia was 14 days (ranged from 1 to 83d). 61 out of 71 patients (84.5%) suffered from Respiratory infection, and 21 patients (35.2%) suffered from bloodstream infection. Microbiological evidence was got in 55 patients. Among 124 isolated species, Gram-negative microorganism (40.7%) was more commonly observed compared with Gram-positive microorganism (37.3%). The total effective rate of tigecycline was 64.8%. The bacterial clearance rates and bacterial hypothetical clearance were 20.8% and 22.6%, respectively. The media time for temperature come back to nomal was 3 days ( ranged from 1 to 10d). The clinical effective rate was 86.7% when tigecycline was administered for more than 9 days, while just 50.0% when administered for less than 9 days ( P = 0.02) . Patients with bloodstream infection got a worse efficacy than those without bloodstream infection ( 50% vs 67.8%, P=0.038). As a second-line anti-infective therapy, the clinical effective rate of tigecycline (70.8%) was mildly higher than those third-line or fourth-line therapy (61.7%) (P= 0.446)). The most common side effects were mild gastrointestinal symptoms including nausea,vomiting and diarrhea. No lethal adverse events was observed.

Conclusions: ur results demonstrated tigecycline was effective and safety for patients unresponsive to carbapenem with febrile neutropenia. A further study with larger number of cases was needed.

Disclosures

No relevant conflicts of interest to declare.
Granulocyte Transfusion in Febrile Neutropenia
Authors: Aydogan Gonul , Keskindemirci Gonca , Tugcu Deniz , Akici Ferhan , Salcioglu Zafer , Akcay Arzu , Sen Hulya , Gokce Muge , Bayram Cengiz , Aycicek Ali , Aktay Ayaz Nuray
Published: October 2019

Background: Over last 30 years cure rates are increasing among oncology patients. As the therapy intensifies, infectious complications become more common. Neutropenia is defined as neutrophil counts of <500/mm³ and it is one of the most frequent and severe complications of pediatric cancer therapy. In 12-17% of patients with febrile neutropenia, there are proven bacteremia and fungal infections. Pediatric oncology patients have a frequency of 30 morbidity and 1% mortality due to infections. Granulocyte transfusions (GTX) were found to be an effective protective and therapeutic strategy in prolonged neutropenia. GTX had been shown to be limiting the duration and complications of neutropenic period, maximum levels of CRP and duration of antibiotic usage especially after myeloablative therapy.

Materials and methods: In this study that was performed at Kanuni Sultan Süleyman Education and Research Hospital pediatric hematology-oncology unit, 170 granulocyte transfusions applied to 37 high risk febrile neutropenia attack of 23 patients were evalauted retrospectively. Granulocyte transfusions were done when the neutrophile counts were between 0 to 300 /mm³ by taking account the general status, physical examination and cultures of patients. Donors were selected according to blood types and their informed cosent were taken. The donors came to the unit one night before, 8 mg intramuscular dexamethasone and 480 mcg of G-CSF were applied. After 12 hours, granulocytes were taken from the donors by intermittent flow technique and given to patients intraveneously. The time needed for getting away from neutropenia, number of febrile days and observed adverse effects were recorded.

Results: For 23 high risk febrile neutropenic patients, granulocyte transfusion was performed at an average of 1-27 times per attack, totally 170 times in 37 attack. Eight of the patients were girls and 15 were boys, ages were ranging between 6 months to 16 years. Product quality qas found to be 1.41 x 1010; 2,14 x 1010/m2. Mean days for subsiding of fever was 6.45 days, mean time for getting rid of neutropenia was 8.15 days. Three patients died during neutropenia attacks. Two of them had both infections and uncontrolled progressive primary disease. Only one patient was lost from infection even though he had antibiotic, antifungal and supportive therapy. Only upper respiratory disease symptoms were noted at 3 of the donors. In one of the patients, transfusion was stopped due to allergic reactions. No other adverse reactions were noted.

Conclusion: Our results were similar to the literature and it shows that granulocyte transfusions after myeloablative therapy limits the neutopenic period, decrease complications of neutropenia, maximum level of CRP and the duration and number of antibiotic usage. But stil controlled studies done at larger cohorts are needed.

Disclosures

No relevant conflicts of interest to declare.
Quantification of Marrow Plasmacytosis in HIV Patients
Authors: O'Brien Timothy , Bowman Lynda
Published: October 2019

Introduction:

A reactive, polyclonal increase in marrow plasma cells in pts with HIV is relatively common, reported to occur in 25% of cases in one series (1) and recently noted to be "usually seen" (2). However, quantification of the degree of marrow plasmacytosis has not been well described.

Methods:

Pts with a diagnosis of HIV and who had a bone marrow aspirate and biopsy performed at our institution over a 10 year period (2/2004-9/2014) were reviewed. Pts with a diagnosis of myeloma were excluded. A 500 cell count was performed on each marrow aspirate and plasma cell percentage determined.

Results:

65 pts were identified; 9 were excluded due to dry taps, so there were 56 evaluable marrow aspirates. Reasons for performing the aspirate and biopsy included: lymphoma staging (NHL in 11/65, HL in 4/65); abnormal SPEP (2/65); abnormal MRI (1/65); FUO (14/65), Kaposi's sarcoma/fevers (3/65), pancytopenia (15/65), thrombocytopenia (6/65), neutropenia (2/65) and anemia (10/65). The average marrow plasma cell percentage was 4.6%, with a range from 0- 21% (median 4%). Only 7/56 (12.5%) had >10% plasma cells. Of these 7 cases, only one had an identifiable cause (multicentric Castleman's) for the plasmacytosis. The 6 others had no neoplastic or infectious etiology, except for HIV itself (including one with 21% plasma cells). All had polyclonal plasmacytosis.

Conclusions:

In this small series of HIV marrows the degree of plasmacytosis was generally mild (average of 4.6%, which is barely above the upper limit of normal range of 3.5%). However, there were a minority of cases which exhibited significant plasmacytosis (as high as 21%) without any cause other than HIV. This suggests that, while significant marrow polyclonal plasmacytosis is usually not seen in HIV, it may occur as a result of HIV itself without any other identifiable cause.

1) Karchen DS and Frost AR. The bone marrow in HIV-related disease: Morphology and clinical correlation. Am J Clin Path. 1991; 95(1): 63-71.

2) Leibman HA and Tulpule A. "Hematologic manifestations of HIV/AIDS". Chapter 159. Hematology: Basic Principles and Practice, 6th edition. 2013.

Disclosures

No relevant conflicts of interest to declare.
First Report from the Saudi Bleeding Screening Study: Are Platelet Disorders More Common in Arabs?
Authors: Owaidah Tarek , Al-Momen Abdul Kareem M , Alzahrani Hazza A , Albanyan Abdulmajeed , Alsulaiman Ayman , Khawar Siddiqui , Alzahrani ALI , al-Madani Mohammad , AlSaleh Khalid , Mahsin Saleh
Published: October 2019

Inherited platelet disorders (IPDs) are a heterogeneous group of diseases affecting platelet production and function with variable clinical severity. The prevalence is estimated to be less than 1:1000,000 worldwide in most common types. There are no population based epidemiological studies in Arab ethnicity to estimate the prevalence of IPD.

Over 2 years we had conducted questioner-based survey among high school and 1st year university students in Riyadh city, Saudi Arabia. We had interviewed 3881 students (1901 (49%) male and 1980 (51%) female). Out of these students 1849(47.6%) had answered yes for any of the questions in first questioner which was followed by a detailed questioner based on MCMDM1 - VWD. Only 323 agreed to have blood testing to investigate for the bleeding tendency. Each positive case was tested for CBC, PT, PTT, ABO and PFA 100 according to standard technique. Any positive screening test for coagulation or platelet screening test was followed by more testing for relevant coagulation factor level. Those were positive only for PFA100 had been tested for platelet aggregation, electronmicroscopy, PB morphology and a sample for DNA and proteomics been collected.

The most common bleeding symptom was epistaxis found in 19.7% (3881) of all interviewed students students and was more in the positive cases 229 (43.6%) students from a total of 525 with more boys (147 out of 296, 49.7%) than the girls (82 out of 229, 35.8%, P-value= 0.002). The cutaneous symptoms were reported by 29.3% (154/525), more common in girls (102/229, 44.5%) than boys (52, 296, 17.6%, p-value<0.001). Out of the investigated cases,( 47 out of 323, 14.5 %) had high PFA100-EPI and (57 out of 323, 17.6%) Had high PFA100-ADP and 22 of these had both cartilage prolonged. We had tested 37 of the suspected cases for platelet disorders for platelet aggregation, only one case had typical features of Glanzmann's disease and 20 cases were abnormal with no clear pattern for known platelet disorders and were labeled as probable platelet disorders. We had diagnosed 8 cases with vWD.

Conclusion: The early results from our bleeding survey indicate that platelet disorders probably more common in our population than been reported els where. We are waiting the final result of laboratory tests which includes ultrastructural features, proteomics and genotyping of these cases with probable platelet diorders

Disclosures

Owaidah: King abdulaziz city for science, Novo Nordisk, Bayer: Honoraria, Research Funding.
Platelet Features from ITP Patients Responders to Different Therapeutic Treatments
Authors: Butta Nora , Rivas Pollmar María Isabel , Álvarez Román María Teresa , Martín Salces Monica , Fernandez Bello Ihosvany , Canales Miguel , Jiménez Yuste Victor
Published: October 2019

Background: Patients with ITP have a wide variation in the presentation of the disease, platelet count and their clinical course.

The decision to begin treatment is based on the hemorrhagic symptoms and platelet count. Intravenous immunoglobulin (IVIG) is usually associated with glucocorticoid administration in patients with severe bleeding or platelet counts <20x109/L and a quick response is required. Agonists of thrombopoietin receptor (TPO-AR) and splenectomy are other therapeutic tools for these patients.

Materials and Methods: We recruited patients with ITP before and after responding to treatment with IVIG (n = 11) and AR-TPO (4 patients with romiplostim and 10 with eltrombopag), 5 splenectomized patients and 82 healthy controls.

The percentage of reticulated platelets, platelet activation and binding of annexin-V were evaluated by flow cytometry. Plasma levels of TPO and "a proliferation-inducing ligand" (APRIL) were determined by ELISA. Procoagulant activity associated microparticles (MP) and the ability of plasma to generate thrombin were determined, respectively, with Zymuphen kit and calibrated automated thrombinography (CAT) triggered by 1 pM tissue factor and 4 micromolar phospholipid (PPP-low reagent, Diagnostica Stago, Spain).

Results: Patients with ITP that respond to IGIV and AR-TPO treatments recovered platelet counts without reaching the levels of the control group, whereas the platelet count in splenectomized patients did not differ from it. Plasma levels of TPO and the number of immature platelets in the first two groups were higher than in controls before responding to treatment. Despite recovering platelet count, platelet capacity of being activated by agonists such as TRAP (thrombin receptor agonist for PAR-1) was less than that of the controls in all groups. This decrease was not due to a reduction in the expression of the fibrinogen receptor on platelets from ITP patients.

Platelets from ITP patients before and after responding to all treatments studied, showed more phosphatidylserine exposure and greater microparticles-associated and plasma-associated procoagulant activity.

Plasma levels of APRIL, a factor that stimulates B cells and antibody production, decreased in ITP patients who responded to the AR-TPO, reaching the levels observed in the control group. In the group of splenectomized patients a decrease of APRIL was also observed, but still remained higher than in healthy controls.

Conclusions: ITP patients who respond to treatment with IVIG and AR-TPO and undergoing splenectomy recovered platelet count but not its function. The treatments did not modify the microparticles- and plasma-associated thrombogenic capacity. Among all the treatments studied, AR-TPO and splenectomy had an addittional benefical effect reducing APRIL plasma levels

Disclosures

No relevant conflicts of interest to declare.
Mod-5014, a Long-Acting FVIIa-CTP, Proposing a Novel Prophylactic Treatment Supporting Less Frequent Subcutaneous or Intravenous Injections with a Similar Mechanism of Action to rFVIIa: Proof-of-Concept in Hemophilic Animal Models
Authors: Binder Lior , Bar-Ilan Ahuva , Hoffman Malka , Hart Gili
Published: October 2019

Introduction:

OPKO Biologics is a clinical-stage public company developing bio-better long-acting versions of existing therapeutic proteins, utilizing a technology termed CTP. The technology involves fusion of the C-terminal peptide of hCG to a target protein. The aim of this work was to comprehensively assess the feasibility of intravenous (IV) or subcutaneous (SC) administration of FVIIa-CTP (MOD-5014) utilizing the most relevant in vivo pre-clinical models, and to characterize the FVIIa-CTP mechanism of action in preparation for an on-going clinical study.

Methods:

FVII-CTP was expressed in CHO cells, purified and activated utilizing a CTP-specific purification process. FVIIa-CTP's pharmacokinetics (PK), pharmacodynamics (PD), long-term hemostatic effect and safety parameters were extensively characterized following SC and IV administration in transient FVII-/- rats and FVIII-/- mice. In addition, the long-term hemostatic effect of FVIIa-CTP was evaluated following a bleeding challenge and compared to commercial rFVIIa. Finally, interaction with co-factors, activity, and the off-target effect of FVIIa-CTP was comprehensively characterized.

Results:

The studies demonstrated that FVIIa-CTP provides long-term exposure (AUC) and half-life that are significantly superior to those of rFVIIa, and consistent with the prolonged half-life of FVIIa-CTP (at an average of 3- and 5-fold, respectively) when compared to IV or SC administration of FVIIa. In addition, a 30% increase in bioavailability was observed relative to commercial FVIIa. A profound improvement in clotting parameters and survival rate following TVT, as well as a reduction of bleeding duration and intensity in tail-clip studies were obtained for both routes of administration for up to 48 hours. Moreover, the safety profile of FVIIa-CTP was further confirmed.

Conclusion:

Attachments of CTP to FVIIa led to a pronounced enhancement of PK and PD, increased exposure as reflected by AUC, elevated half-life, and improved recovery in mice, rats and pigs following SC and IV administration. FVIIa-CTP injection resulted in an improved bioavailability that translated to a marked in vivo hemostatic effect. Our data suggest that CTP-fused FVIIa can potentially provide a novel approach for IV or SC prophylactic treatment of hemophilic patients (both pediatric and adult), with the major benefit of significant improvement in quality of life.

Disclosures

No relevant conflicts of interest to declare.
Evaluation of Triton X-100 Effect on Coagulation Tests
Authors: Nadeau Marchand Sarah , Khalaf Daniel , Nadeau Chantal , Blais Normand
Published: October 2019

CONTEXT:

Triton X-100 is a non-ionic surfactant that has been proposed as a virus inactivator in a laboratory setting for it causes cell lysis through lipid membrane disruption without denaturing proteins. Very few studies have tried to evaluate its effect on laboratory tests, especially on coagulation tests that require a phospholipid subtract.

METHOD:

Plasma samples were collected. Triton X-100 was spiked at a concentration of 0,25% and specimens were incubated for an hour. We performed international normalized ratio (INR), activated partial thromboplastin time (aPTT), thrombin time (TT), fibrinogen dosing and D-Dimer dosing, comparing the standard plasma with the triton X-100 exposed plasma. We also dosed coagulation factors, and performed dilution and lupus anticoagulant studies. Finally, we compared prothrombin time and aPTT using reagents from various manufacturers.

RESULTS:

We observed an important difference for the INR and the aPTT with addition of triton x-100. A slight decrease of an average of 6.6 to 16.4% was also found in factor assays. Differences in the INR and the aPTT were dependent respectively on the ISI and the phospholipids content of the reagent.

CONCLUSION:

The addition of triton x-100 significantly modifies INR and aPTT results with every reagent tested. The impact on TT and fibrinogen is not clinically relevant, and the D- Dimer dosage is not affected. The slight decrease in all coagulation factors does not explain those results. The interference could result from an interaction with phospholipids, calcium or some enzymatic reactions caused by the presence of triton x-100 in the analyzed specimen.

Table. Test Mean Mean Mean of difference p-value (paired t-test) without Triton with Triton INR 1,59 2,87 68,80% p<0,001 aPTT 31,59 sec 61,80 sec 86,10% p<0,001 TT 16,82 sec 17,42 sec 3,50% p<0,001 Fibrinogen 3,27 g/L 3,21 g/L -0,70% p=0,001 D-Dimer 2897 mcg/L 2905 mcg/L 4,30% p=0,697

Disclosures

No relevant conflicts of interest to declare.
Carbohydrate Glycosylation Deficiency Disorder Secondary to Absence of Phosphomannose Mutase Confers Both a Hemorrhagic and Thrombotic Phenotype
Authors: Gleditsch Katrina , Ameri Afshin
Published: October 2019

Congenital disorders of glycosylation are a group of inherited disorders affecting the addition of a carbohydrate moiety to a protein. A defect in this pathway can cause adverse effects in most organ systems, often presenting early in life. The most common of these disorders is CDG type 1a / PMM2-CDG). In this disorder the loss of the enzyme phosphomannomutase-2 prevents the conversion of mannose-6-phosphate to mannose-1-phosphate. This has previously been associated with deficiencies of protein C, protein S, antithrombin III, factor IX and factor XI and a subsequent imbalance of coagulation pathways leading to thrombotic events.

We present the case of a 14-year-old male with known history of CDG1a and previous bleeding complications following left orchiopexy at age 3, at that time coagulation screening showed a normal PT/PTT and surgery was deemed safe. The patient now presented to our pediatric hematology/oncology clinic for further evaluation of coagulation abnormalities prior to pediatric surgery performing right orchiopexy. Laboratory values at this time showed coagulation deficiencies, of ATIII, Factor XI, Protein C and Protein S, PT/ PTT , F VIII, FIX and vWF where normal. Prior to surgery, patient was given fresh frozen plasma and ATIII concentrate the patient underwent a successful Stage One Fowler-Stephen Procedure with adequate hemostasis. Several months later the patient developed leg swelling and was diagnosed right femoral DVT. This resolved with ATIII substitution and anticoagulation with LMW heparin. The patient's immediate family was tested for bleeding/clotting disorders and results were found to be normal. This report not only supports the association of bleeding but also thrombosis with CDG1a. We propose that in patients with known CDG routine PT/PTT will not uncover a hemostatic abnormality but further screening isolated factor deficiency need to be performed and prophylactic factor substitution be performed prior to any surgical interventions. Also an awareness of the highly procoagulant state in these patients that predispose to DVT and central nervous system vascular thrombosis need to be present.

Disclosures

No relevant conflicts of interest to declare.
Immune Hemolysis and Aseptic Meningitis after Intravenous Immunoglobulin
Authors: Arumugam Raajasekar Arun Kumar , Nannapaneni Srikant , Solomon William B.
Published: October 2019

INTRODUCTION

Human intravenous immunoglobulin (IVIG) products are generally considered safe and are used for the treatment of a variety of autoimmune and pro-inflammatory states. However, controlled trials involving IVIG have been of small size with limited power and described only the most common adverse effects [1]. IVIG related hemolysis is a rare adverse effect and is usually self-limited, rarely requiring blood transfusions. A few case reports have also described aseptic meningitis after IVIG use. We describe a patient who developed severe hemolytic anemia and aseptic meningitis after IVIG use.

CASE PRESENTATION

A 54 year old woman was admitted with severe head ache, dark colored urine and shortness of breath twenty four hours after receiving IVIG. She was diagnosed with chronic inflammatory demyelinating polyneuropathy (CIDP) a month ago and was started on IVIG. Physical exam was unremarkable. Laboratory tests showed severe anemia with a hemoglobin of 6.1 g/dl (baseline 13 g/dl), indirect hyper-bilirubinemia, low haptoglobin, elevated LDH and hemoglobinuria confirming hemolysis. Hematological testing revealed a positive direct antiglobulin test (DAT), DAT IgG, DAT CD3 and eluate testing were also positive. A lumbar puncture showed neutrophil pleocytosis but no bacteria. A diagnosis of IVIG induced hemolysis and aseptic meningitis was made. O negative packed red blood cells were transfused for severe symptomatic anemia and non-steroidal anti-inflammatory drugs were given for headache. Steroids were deferred since the source of antibodies causing hemolysis was extrinsic. She improved clinically and blood counts returned to baseline in three weeks.

DISCUSSION

Pooled IVIG is extracted from at least 1000 individuals and contains highly purified polyvalent IgG [2]. Hemolysis after IVIG is thought to be secondary to donor antibodies against host red blood cell antigens and resolves once IVIG is stopped. Risk factors for hemolysis include non-O blood types and the use of high dose IVIG. The patient described was of AB Rh positive blood type but did not receive high dose IVIG. Aseptic meningitis is another rare adverse effect that has been postulated to be related to antibodies in the IVIG that mimic antineutophil cytoplasmic antibodies (ANCA) which activate neutrophils causing neutrophilic pleocytosis [3]. One must be aware of these two rare complications of a commonly used drug.

REFERENCES

[1] Risks associated with the use of intravenous immunoglobulin. Pierce LR et al. Transfus Med Rev. 2003 Oct;17(4):241-51.

[2] Appropriate uses of human immunoglobulin in clinical practice: memorandum from an IUIS/WHO meeting. Bull World Health Organ. 1982;60(1):43.

[3] Intravenous immunoglobulins contain naturally occurring antibodies that mimic antineutrophil cytoplasmic antibodies and activate neutrophils in a TNF alpha-dependent and Fc-receptor-independent way. Jarius et al. Blood. 2007;109(10):4376.

Disclosures

No relevant conflicts of interest to declare.
Presence of Primary Cilia in Human Hematopoietic System
Authors: Singh Mohan , Chaudhry Parvesh , Ramsingh Giridharan , Merchant Akil
Published: October 2019

Primary cilia are non-motile microtubule-based organelles that are present on the cellular membrane of all eukaryotic cells. The presence of cilia is required for the response to developmental signaling pathways such as Hedgehog (Hh) and Wnt/β-catenin. Loss of primary cilia or their function has been linked to a number of human genetic diseases, called ciliopathies (Dorn et al., Dev Cell, 2012). Recent studies suggest that abnormal hedgehog Hh pathway activity due to ciliary dysfunction could lead to various cancers such as glioblastoma, medulloblastoma, basal cell carcinoma, breast, prostate and ovarian cancer (Menzl et al., Cilia, 2014, Moser et al., BMC cancer 2009). Although the Hh pathway has emerged as an exciting new therapeutic target in leukemia and myeloproliferative diseases, to date, there have been no reports describing the presence of primary cilia in human blood or leukemia cells. In the present study, we have successfully identified primary cilia in hematopoietic system (Fig. 1). Primary cilia are well characterized in hedgehog responsive NIH3T3 cells; therefore, these cells were included in the study as a positive control for cilia staining. Localization of acetylated tubulin (which stains for the ciliary body) and gamma tubulin (which stains the basal body) was used to confirm the presence of primary cilia in normal human peripheral blood mononuclear cells (PBMCs) and normal bone marrow (BM) cells. Furthermore, we have also identified primary cilia in acute myeloid leukemia cell lines (KG1, KG1a, K562). Additionally, hedgehog pathway components such as, Smoothened and Gli3 were localized in primary cilia of acute myeloid leukemia (AML) cell lines. Interestingly, the frequency of primary cilia in AML cells (10-36%) was found to be lower than normal PBMCs and BM (97-99%) (Fig. 1). Moreover, cilia present on leukemic cells were often shorter and dysmorphic, with less well defined basal bodies. KG1 cells, which have a higher number of cilia, retain sensitivity to Hh pathway inhibitors, but K562 and KG1a, which have high levels of Hh activity but very low numbers of cilia, are resistant. Loss of cilia is, therefore, associated with activation of Hh signaling that is resistant to SMO antagonists, and could have important implications for the development of this class of drugs. In conclusion, we present the first evidence showing the presence of primary cilia in hematologic system and implicate the loss of primary cilia in development of hematological malignancies through deregulated Hh signaling.

Disclosures

No relevant conflicts of interest to declare.
Abnormal Levels of PKM2 in Mdcs from Patients with Severe Aplastic
Authors: Shao Zonghong , Zheng Mengying
Published: October 2019

Severe aplastic anemia (SAA) is a hematologic disease characterized by pancytopenia with severe bone marrow failure. Our previous studies have demonstrated that activated myeloid dendritic cells (mDCs) increased in the bone marrow of SAA patients, which might promote Th0 cells to polarize to Th1 cells and cause the subsequent over-function of T lymphocytes and hematopoiesis failure in SAA. The other notable finding in our study is that abnormal expression of pyruvate kinase M2 (PKM2) in mDCs from SAA patients may be one of the reason for mDC hyperfunction. Human pyruvate kinase M2, which catalyzes the last but rate-limiting step of glycolysis, is exclusively expressed in actively proliferated cells ,especially in embryonic and adult dividing/tumor cells. It remains unclear how PKM2 may change in patients with SAA. This study aims at exploring the role of PKM2 in SAA. In this study, the quantitative changes of mDCs in peripheral blood of 15 SAA patients before immunosuppressive therapy (IST) °¢ 13 SAA patients after IST and 26 normal controls were determined by flow cytometry. Results showed that the levels of PKM2 in mDCs were significantly increased in SAA patients(59.1±15.8)%. After IST, the levels of PKM2 in mDCs decreased(42.6±22.1)% (P<0.05).In normal controls group , the level of PKM2 in mDCs was (32.7±20.2)%. These changes reveal that dysregulation of PKM2 expression and activation in mDCs from SAA patients may contribute to hyperfunction of mDCs, thus leading to the imbalance of downstream Th1/Th2 subsets which followed by hematopoiesis failure in SAA.

Figure 1. Figure 1.

Disclosures

No relevant conflicts of interest to declare.
Tumor Microenvironment Regulates Survival of Mantle Cell Lymphoma Cells through Various Signaling Pathways
Authors: Sehgal Lalit , Tamer Khashab , Wang Xin , Mathur Rohit , Berkova Zuzana , Romaguera Jorge E. , Rodriguez Alma , Neelapu Sattva S. , Samaniego Felipe
Published: October 2019

Introduction

An aggressive, incurable form of non-Hodgkin's lymphoma (NHL) is classifed as Mantle cell lymphoma (MCL). The health complications associated with advanced age of restrict treatment with intense chemotherapy. Overexpression of cyclin-D1 due to Translocation t(11;14) , is the hallmark of MCL. More detailed insight into MCL pathogenesis has been delayed until the recent development of a tissue culture system, using human mesenchymal stromal cells (hMSC), suitable for propagating primary MCL cells. We hypothesized that tumor-initiating cells are responsible for MCL relapse and chemoresistance and thus, identification of signals responsible for survival and maintenance of MCL-initiating cells (MCL-ICs) is essential for design of curative treatment strategies.

Methods

Isolates of primary MCL cells (n=24) were co-cultured with human mesenchymal stem cells (hMSCs) and the content of MCL-ICs was analyzed by flow-cytometry based on marker expression profile; CD34-CD3-CD45+CD19-. Cytokine array was used to identify the soluble factors enriched in the co-cultures and the expression of these factors was confirmed by RT-PCR analysis. The signaling pathways employed by the newly-identified factors were blocked in 3 MCL cell lines (JVM2, Mino, Z138) to confirm their essential role in survival of MCL cells and, more importantly, for MCL-ICs.

Results

Co-cultures of primary MCL isolates with hMSCs supported the growth of MCL cells for over 4 weeks with continued presence of MCL-ICs (CD34-CD3-CD45+CD19-) representing about 1% of MCL cells. We found that IL-6 produced by hMSCs triggered an FGF/FGFR autocrine loop in MCL-ICs. The extent of FGFR expression correlated tightly with expression of SOX11, a pathology related negative prognostic marker in MCL. MCL cell survival and growth was regulated via the FGFR/mir101/ EZH2/ NF-κB/XIAP axis. Blocking of this signaling pathway with FGFR1 inhibitors consistently induced early reduction in XIAP levels and subsequently MCL cell death.

Conclusion

We established that propagation of primary MCL in co-cultures with hMSCs depends on an FGF/FGFR autocrine loop that enhances XIAP protein expression and thus, supports survival of MCL cells. We identified the factors essential for survival of MCL and MCL-ICs that present new targets for improved MCL treatment strategies.

Disclosures

Rodriguez: Orthobiotech: Research Funding.
Improved Molecular Diagnosis of Aplastic Anemia/Bone Marrow Failure Syndromes Using a Novel Comprehensive Next Generation Sequencing Gene Panel
Authors: Dasouki Majed , Ahmed Syed Osman , Alahmari Ali , Jabr Amal , Alawwami Moheeb Ali , Chaudhri Naeem A. , Fahad AlMohareb , Mohamed Said Y , Rasheed Walid , Alzahrani Hazza A , Aljurf Mahmoud
Published: October 2019

Introduction: Various forms of aplastic anemia (AA)/bone marrow failure syndromes (IBMFs) show significant clinical and molecular heterogeneity with significant clinical overlap and are often diagnosed based on established clinical and pathological criteria. While >70 genes have been identified in patients with AA/IBMFs, most cases have are labelled as idiopathic with no identifiable genetic abnormlaity found. Precise detection of genetic abnormalities in these patients may assist in more accurate molecular diagnosis in these patients, proper counseling, cancer surveillance and personalized clinical intervention.

Method: As part of the Saudi Human Genome Project, we developed a comprehensive 405 gene panel encompassing all known Mendelian hematological disorders (hemolytic anemias, aplastic anemias/bone marrow failure syndromes, coagulation disorders) using the Ion Torrent AmpliSeq technology. Patients who met the clinical diagnosis of aplastic anemia/bone marrow failure syndrome were enrolled into this study. Peripheral blood samples were subjected to this next-generation sequencing analysis.

Results: We validated the Saudi Mendeliome assay using 642 samples with known mutations across various medical specialties. We then tested 37 patients with AA/IBMFS using this Proton-Ion sequencing platform. Mutations were identified in 7/37 (19%) of patients, followed by whole exome sequencing (WES) in those patients without identifiable mutations.

Conclusion: Compared with clinical WES and/or whole genome sequencing (WGS), which are still expensive, time consuming and difficult to interpret, this novel and comprehensive targeted gene panel is more economical (< $150), faster (3-4 weeks), upgradable (by spiking in newly identified AA/IBMFs genes) and can be used to genotype patients with acquired aplastic anemia/bone marrow failure syndromes and guide their management. Second tier testing using WES/WGS is recommended for cases without identifiable mutations.

Disclosures

No relevant conflicts of interest to declare.
A Small-Molecule Suppressant of Survivin YM155 Induces Cell Death Via Proteasomal Degradation of c-Myc in Multiple Myeloma Cells
Authors: Ito Shigeki , Asahi Maki , Ishida Yoji
Published: October 2019

[Background] Survivin is a member of the inhibitor of apoptosis protein (IAP) family with its dual roles in mitosis and apoptosis, and emerges as an attractive target for cancer therapy. YM155, a novel molecular targeted agent, suppresses survivin, which is overexpressed in many tumor types. However, the effect of this agent on multiple myeloma (MM) cells remains unclear. [Materials & Methods] Five human MM cell lines, RPMI8226, U266, KMS20, KMS28PE, and KMS34 were used in this study. Cell proliferation and cell death were evaluated by MTT assay and by flow cytometric analysis with annexin V/PI staining. Gene and protein expressions were analyzed with quantitative PCR (qPCR) and immunoblot, respectively. For proteasome inhibitory assay, cells were treated with YM155 and/or proteasome inhibitor MG132 for 6 hours. [Results] YM155 inhibited cell proliferation of these cells in a dose- and time-dependent manner. Annexin V assay showed that YM155 induced apoptosis in these cells. To better understand these effects of YM155 on MM cells, we evaluated the intracellular signaling and apoptosis-associated protein status. Immunoblot analyses showed that YM155 reduced not only survivin but also Mcl-1 and XIAP expressions. We also observed the activation of caspase-3 and PARP in YM155-treated cells, indicating that YM155 induces caspase-dependent apoptosis. In contrast, YM155 did not affect phosphorylation status of Erk1/2 and STAT3. Interestingly, YM155 suppressed c-Myc and IRF4 protein expressions, both of which are recognized as an important transcription factor in the pathogenesis of MM. In addition, qPCR assay showed that YM155 treatment did not reduce c-Myc mRNA level. On the other hand, proteasome inhibitor prevented the suppression of c-Myc expression by YM155 treatment, indicating a proteasomal degradation of c-Myc by YM155. [Conclusion] YM155 suppresses cell proliferation and survival in MM cells in part via not only inhibiting anti-apoptotic proteins such as survivin, Mcl-1 and XIAP but also suppressing c-Myc oncoprotein expression. Further study is needed to clarify the molecular mechanism of c-Myc degradation induced by YM155. Our results may provide a new concept in c-Myc-targeting therapy for MM because c-Myc oncoprotein have been considered undruggable.

Disclosures

Ishida: Bristol-Myers Squibb: Honoraria.
Cross Talk Between Haemostasis and Oxidative Stress in Acute Leukemia
Authors: Naz Arshi , Sattar Abdul , Farzana Tasneem , Taj Mehwish , Ansari Saqib Hussain , Nadeem Mohammad , Shamsi Tahir
Published: October 2019

Objective: To investigate the haemostatic/oxidative changes and its association in patients of acute leukemia at the time of diagnosis.

Study Design: Cross sectional and descriptive.

Setting: Study was carried out in National Institute of Blood Disease and Bone Marrow Transplantation during 2011 to 2013.

Material and Methods: Activity of Superoxide dismutase(SOD), Glutathione Reductase(GR), Total antioxidant status(TAS), Prothrombin Time, Activated Partial Thromboplastine Time, Fibrinogen, Factor VIII and Tissue Factor were evaluated in the hemolysate, sera, citrated plasma and leukemic cells of 150 patients with acute leukemia at the time of diagnosis in 40 healthy individuals.

Results: The mean age of patients was 29.8 years.Activity of SOD, GR and TAS were significantly higher in patients of acute leukemia as compared to control (<0.01). Significant haemostatic changes were identified in patients of acute leukemia. Strong association was found between oxidative stress/total antioxidant status markers and haemostatic parameters which are in favor of cross talk between haemostasis and oxidative stress in acute leukemia.

Disclosures

No relevant conflicts of interest to declare.
A New Strategy to Target Acute Myeloid Leukemia Stem and Progenitor Cells Using Chidamide, a Histone Deacetylase Inhibitor
Authors: Xu Bing , Li Yin , Chen Kai , Zhou Yong , Xiao Yiren , Deng Manman , Jiang Zhiwu , Ye Wei , Wei Xinru , Wang Xiangmeng , Li Jie , Liang Jiabao , Zheng Zhongxin , Yao Yao , Wang Weiguang , Li Peng
Published: October 2019

Leukemia stem cells (LSCs) are responsible for treatment failure and relapse in acute myeloid leukemia (AML). Therefore, development of novel LSCs-targeting therapeutic strategies is of crucial clinical importance to improve the treatment outcomes of AML. Histone deacetylase (HDAC) inhibitors have emerged as a potential strategy to reverse aberrant epigenetic changes associated with cancer. HDAC inhibitors have shown potent and specific anticancer stem cell activities in preclinical studies. Chidamide, a novel benzamide-type selectively HDAC inhibitor, has been reported to induce G1 arrest and apoptosis in the relatively mature progenitor population, whereas its effect on primitive LSCs has not been clarified. In this study, we demonstrated that chidamide specifically induces apoptosis in LSC-like cells and primary AML CD34+ cells in a concentration- and time-dependent manner. Our further molecular mechanistic study uncovered that chidamide induces LSCs death by activation of reactive oxygen species (ROS). It compromises the mitochondria membrane potential, modulates anti-apoptotic and pro-apoptotic proteins in BCL2 family and activates caspase-3 leading to PARP degradation. Meanwhile, chidamide activates CD40 and modulates its downstream signaling pathways, JNK and NFκB. The results of this study suggest that chidamide may be a novel LSC-targeting agent for AML therapeutics.

Disclosures

No relevant conflicts of interest to declare.
Morphologic Evolution in Post-Transplant Lymphoproliferative Disorders (PTLD): A Clinicopathologic Case Series
Authors: Stubbins Ryan J. , Peters Anthea C , Curtis Mabilang , Owen Carolyn , Mansoor Adnan , Lai Raymond , Preiksaitis Jutta
Published: October 2019

Post-transplant lymphoproliferative disorders (PTLDs) are classified as early PTLDs (E-PTLD), polymorphic PTLDs (P-PTLD), monomorphic PTLDs (M-PTLD) and classical Hodgkin's Lymphoma PTLD (HL). These entities are felt to represent a disease continuum, with a precursor-product relationship, though they are morphologically and clinically distinct. However, this process remains poorly understood, and limited evidence exists to support this hypothesis. We report a series of nine cases extracted from a PTLD database, including both pediatric and adult solid organ transplant recipients, with recurrent disease episodes that evince an apparent evolution in their morphologic categorization between episodes. All patients identified were high risk for PTLD, with multiple identifiable predisposing factors. Presentations varied from isolated lymphadenopathy, to gastrointestinal involvement to pulmonary involvement. Four of the patients were deceased at the time of acquisition, though only one directly from PTLD. Eight of the nine identified patients developed E or P-PTLD lesions that preceded a subsequent M-PTLD or HL lesion at a similar tissue site. Two of the cases had metachronous P and M-PTLD lesions. The six M-PTLD subtypes were variable, including DLBCL, Burkitt, T-cell, and extramedullary plasmacytoma, in addition to the three cases of classical Hodgkin's lymphoma. These cases suggest that E and P-PTLD may act as common precursor lesions to the development of both the M-PTLD variants and HL type lesions, and that the PTLD classification schema represents different stages of a common underlying pathology.

Disclosures

Owen: Lundbeck: Honoraria; Gilead: Honoraria; Janssen: Honoraria; Roche: Honoraria.
Multiple Clonal TCR Gene Rearrangements Are Typical in Peripheral T-Cell Lymphoma Not Otherwise Specified
Authors: Sidorova Yulia V , Chernova Natalia G. , Ryzhikova Natalia V. , Smirnova Svetlana Yu. , Vinogradova Yulia E , Kovrigina Alla M. , Zvonkov Eugene E. , Sudarikov Andrey B. , Parovichnikova Elena N. , Savchenko Valeri G.
Published: October 2019

Introduction: Peripheral T-cell lymphoma not otherwise specified (PTCL-NOS) is a heterogeneous subgroup of PTCL with no consistent immunophenotypic, genetic and clinical features. PTCL-NOS represents less than 29% of all T-cell lymphomas in adults and often has an aggressive course with extranodal and systemic involvement. This study is focused on the application of clonality testing methods for PTCL-NOS diagnosis and monitoring.

Patients and Methods: Biopsies, blood and bone marrow samples were tested for 30 patients with PTCL-NOS: the ratio of male/female was 15/15, median age was 56 years (32-75), and 22 of 30 patients had IV stage of the disease. TCRG and TCRB gene rearrangements were evaluated for T cell clonality analysis. We used PCR-based methods (BIOMED-2 standardized clonality protocol) followed by GeneScan analysis on the ABI PRISM 3130 Genetic Analyzer (Applied Biosystems).

Results: Clonal TCR gene rearrangements were found in 29 of 30 patients (97%): TCRG in 27 of 30 (90%) and TCRB in 29 of 30 (97%). Primary biopsies or bone marrow samples showed oligoclonal pattern of rearrangements (more than 3 clonal peaks in TCRG or TCRB GeneScan profiles) in 13 of 30 patients (43%). New clonal rearrangements in affected tissues (skin, tonsil, CSF, lymph nodes and etc.) during disease progression were found in 15 of 24 (63%) patients. Various representations of clonal rearrangements in different tissues indicate the presence of several tumor clones in the majority of cases (19 of 30 patients; 63%). The presence of several clones correlated with the number of tissues examined (rs = 0.6, p <0.0005). The detection rate of additional clonal rearrangements was 90% (9 of 10 patients) when 3 and more tissues were tested. The number of clonal rearrangements was not correlated with age (p = 0.43) or the stage of disease (p = 0.29). One can speculate that the presence of multiple clones in the PTCL-NOS could reflect genetic instability of this tumor.

Conclusion: Oligoclonality, clonal evolution and the appearance of new clones in the progression of the disease are typical features of PTCL-NOS. Multiple clonal rearrangements complicate diagnostics and assessment of minimal residual disease in this disease.

Disclosures

No relevant conflicts of interest to declare.
Outcome of Elderly Patients with Hodgkin's Disease-Single Centre Experience from Riyadh, Saudi Arabia
Authors: Tailor Imran Khan , Zaidi Syed Ziauddin A. , Motabi Ibraheem H , Alshehry N F , Elghazaly Assem A , Almudaibigh Samer Homoud S , AlGhamdi Mubarak S. , Gill Atta Munawar , Iqbal Shahid , Murtaza G , Butt A
Published: October 2019

Background: Hodgkin's disease accounts for approximately 10% of all lymphoma cases. There is a bimodal distribution with initial peak in second decade and a later peak in sixth decade. There is paucity of date on outcome of elderly patients with Hodgkin's disease. There is hardly any data from middle eastern patients.

Materials and Methods: We retrospectively analyzed 168 consecutive patients diagnosed with Hodgkin's disease at our centre from 2006 to 2015 (age>14 years) and found 16 patients to be over the age of 60 years.

Results: Out of 16 patients 6 were females, 10 were males and the median age was 70 years (range 61-86).All patients had classical Hodgkin's disease out of which 12 had nodular sclerosis subtype, 2 had mixed cellularity subtype and 1 had lymphocyte rich subtype. 4 patients had early stage disease (IA-IIA) while rest 12 had advanced stage disease. 15 were treated with ABVD based regimen, the other was treated with chlorambucil based regimen.3 out of 16 were given radiation in addition to chemotherapy. Out of the 14 evaluable patients (two were lost to follow up) 13 completed treatment and all of them achieved complete remission (93%).9 of 16 patients were given reduced dose doxorubicin. Three patients died (one due to sepsis, the other due cardiomyopathy 2 years after treatment, third one due secondary cancer).None of the alive patients have relapsed yet. One patient developed secondary cancer who later died with the same. At a median follow up of 35 months (range 1-72) even free survival and overall survival were 79% and 86% respectively.

Discussion: As per our study unlike in western world, older patients are less frequently affected with Hodgkin's disease. The remission rates and survival data are very encouraging despite dose reductions, however further data from this part of the world is needed to confirm our results. ABVD appears feasible, effective treatment option for elderly patients although frail patients who cannot tolerate ABVD may be candidates for anti CD30 antibody based regimens.

Disclosures

No relevant conflicts of interest to declare.