bluuebunny/crossref_metadata_2025 huggingface dataset:
Cross-Reactivity of Drug-Dependent Antibodies in Patients with Immune Thrombocytopenia Induced By Beta-Lactam Antibiotics
Authors: Slaught Matthew John , Bougie Daniel W. , Aster Richard H.
Published: November 2019

More than 50 beta lactam (BL) antibiotics are now in active use for treatment of a wide range of bacterial infections. BL antibiotics are among the most common drugs capable of inducing antibodies (DDAbs) that cause drug-induced immune thrombocytopenia (DITP). Most DDAbs are highly specific for the sensitizing drug but beta lactams all have a common core structure and many similarities among side groups that are added to augment potency and modify specificity, raising the possibility that a DDAb specific for one BL may cross-react with another. We studied DDAbs from 33 patients with DITP induced by 9 commonly used BL drugs to determine whether patterns of cross-reactivity exist that might influence the choice of an alternative antibiotic in a patient with BL-induced DITP.

DDAbs were demonstrated in a flow cytometric assay considered to be "positive" when immunoglobulins in patient serum but not normal serum react with normal platelets in the presence, but not in the absence of drug (Blood 2018;131:1486). DDAbs detected in the 33 patients were specific for 9 different BL drugs that were divided into two groups, "penicillins" (Group 1) and cephalosporins (Group 2) on the basis of structural similarities (Figure 1). In Group 1 were 19 DDAbs specific for amoxicillin (2), nafcillin (4) and piperacillin (13). Structurally similar ampicillin and penicillin were also tested with these abs. In Group 2 were 14 DDAbs specific for cefadroxil (1), cefepime (2), ceftazidime (2), ceftizoxime (1), ceftriaxone (7) and cephalexin 1). Cross-reactions identified within these groups of DDAbs are shown in Tables 1 and 2.

Cross-reactions, many quite strong (S) were observed among DDAbs specific for drugs in both structural groups (Tables 1 and 2). Particularly noteworthy were cross-reactions of the 19 Group 1 DDAbs with ampicillin (6) and penicillin (6) (Table 1) and of the 14 Group 2 DDAbs with cefepime (6), ceftizoxazole (6) and ceftriaxone (3) (Table 2). The findings show that platelet-specific DDAbs induced by beta lactam antibiotics, in contrast with those induced by medications like quinine, sulfamethoxazole and vancomycin, commonly cross-react with other antibiotics of this class. In patients with immune thrombocytopenia induced by a beta lactam antibiotic, it may be prudent to avoid switching to another beta lactam or, if this is necessary, to monitor platelet counts carefully.

Disclosures

No relevant conflicts of interest to declare.
Combining HPFH Mutations in Human Adult HSCs to Enhance Reactivation of Fetal Hemoglobin
Authors: Psatha Nikoletta , Georgakopoulou Afrodite , Li Chang , Sova Pavel , Wang Hongjie , Lieber Andre , Papayannopoulou Thalia
Published: November 2019

Despite the significant advances in developing novel therapies for hemoglobinopathies, developing a universal and robust therapeutic approach, able to achieve an event-free result in all β0/β0 thalassemic and sickle cell patients is a work in progress. It is known that increase of Fetal hemoglobin (HbF) can lead to an ameliorated clinical picture up to transfusion independency. Inactivation via genome editing of γ-globin suppressors or introduction of Hereditary Persistence of Fetal Hemoglobin (HPFH) mutations in the promoter of hemoglobin gamma (HBG) have been shown to significantly increase the endogenous HbF expression. Here we studied the effect in HbF reactivation by targeting the binding site of the two main HbF silencers, BCL11a and LRF and how it compares to the well-established editing of the BCL11a erythroid enhancer. Furthermore, in order to achieve higher levels of fetal hemoglobin, we assayed the effect of all combinations of these cis and trans acting mutations. We first observed that all single knock outs had similar effects in HbF reactivation. Interestingly, by targeting both sites in the gamma globin promoter we observed a decrease in HbF induction compared to each single edit, suggesting a possible binding of a gamma globin activator in the in-between region. However, editing of the BCL11a erythroid enhancer and either of the two silencer binding sites yielded maximum levels of fetal hemoglobin, with pancellular HbF expression, both in vitro and in vivo following xeno-transplantation in NSGW mice. Collectively, our data suggest that the combination of two fetal globin reactivation mutations, one cis and one trans, have the potential to significantly increase HbF both totally and on per cell basis. This strategy has the potential to induce higher levels of HbF reactivation with a clinical benefit in patients with beta globin disorders.

Disclosures

No relevant conflicts of interest to declare.
Enumerating Functional HSC Clones in the Zebrafish Using Gestalt
Authors: Teets Emily M , Gregory Charles Thomas , Shaffer Jami L , Blachly James S. , Blaser Bradley W
Published: November 2019

Hematopoietic stem cells (HSCs) are functionally and genetically diverse. This diversity decreases as a function of age and with the development of myeloid neoplasia. Numerous systems have been developed to genetically "barcode" HSCs and quantify HSC clone numbers in the setting of native hematopoiesis, but no experimental and computational framework has been established to perform this analysis on a large number of biological replicates. Here we have employed the transgenic zebrafish system, Genome Editing of Synthetic Target Arrays for Lineage Tracing (GESTALT), to develop a framework scaled to permit the investigation of putative oncogenes or niche factors with moderate to small effect sizes. The GESTALT line carries a single copy of an array of 10 tandem CRISPR target sites that is highly variable after being repaired by non-homologous end joining. GESTALT embryos were injected with sgRNAs and Cas9 protein to generate genetic barcodes at the GESTALT locus. Fifty-six zebrafish were raised to adulthood and GESTALT variants were read from a sample of peripheral blood by amplicon sequencing. In a training set of 35 fish we developed a computational model to discriminate informative barcodes from uninformative GESTALT variants in each sample. This model identifies variants shared among samples and excludes them as uninformative. Experimentally-defined thresholds were used to minimize inter-sample correlation (mean Pearson Correlation Coefficient = 0.0021) while maximizing the number of retained informative barcodes in each sample. Bootstrapping and k-means clustering were used to classify samples with a high or low fraction of informative (FI) barcodes (FI cutoff = 0.65). 24 samples within the training set were classified in the High-FI group (mean FI = 0.93 ± 0.02, compared to 0.40 ± 0.04 in the Low-FI group). We then evaluated 3 methods of quantifying GESTALT HSC clones from all sequenced barcodes in the High-FI samples and found that counting barcodes with a VAF>0.02 produced the most normal distribution across the training set (Shapiro-Wilk test for normality, W = 0.95, p = 0.21). 4.3 ± 0.38 GESTALT HSC clones were detected in the training set. In an independent validation set of 21 samples, the number of GESTALT HSC clones was 3.4 ± 0.55, (p = 0.2). This study provides the largest reference dataset to date for HSC genetic barcoding experiments in the zebrafish and addresses the validity of barcode sequences, sample selection and clone enumeration in a systematic, objective way. The GESTALT barcoding system and our analysis framework compare favorably with other methods of studying HSC clonal diversity under conditions of native hematopoiesis in terms of scale, cost, labor and barcode validity. Future studies aiming to understand the mechanisms of HSC clonal evolution in the context of health, ageing and disease will benefit from this framework for enumerating functional HSC clones.

Disclosures

No relevant conflicts of interest to declare.
Inflammatory Cytokine Responsive Enzymatic Mutagenesis Fuels Myeloproliferative Neoplasm Pre-Leukemia Stem Cell Evolution
Authors: Jamieson Catriona , Jiang Qingfei , Holm Frida , Isquith Jane , Mark Adam , Mason Cayla , Ma Wenxue , Diep Raymond H , Pham Jessica , Reynoso Moreno Eduardo , Nasamran Chanond A , Guorong Xu , Birmingham Amanda , Crews Leslie A , Alexandrov Ludmil , Pineda Gabriel , Whisenant Thomas , Fisch Kathleen M
Published: November 2019

Innate immune anti-viral adenosine to inosine (A-to-I) base editing enzymes (editases) promote hematopoietic stem cell (HSC) self-renewal and protect the human genome from retroviral integration in response to inflammatory cytokine signaling. However, hyper-editing has been linked to therapeutic resistance and cancer progression. Because myeloproliferative neoplasm (MPN) progression is typified by increased JAK2/STAT-mediated cytokine signaling, we investigated the cell type and context specific role of adenosine deaminase acting on RNA1 (ADAR1) editaseactivity in MPN pre-leukemia stem cell (pre-LSC) evolution into acute myeloid leukemia stem cells (LSCs). Here we show by whole transcriptome sequencing (RNA-seq) of 113 FACS-purified hematopoietic stem cells and progenitors from 78 individuals, including 54 MPN and AML patients and 24healthy young and aged individuals, that anti-viral signaling pathway activation and splice isoform switching from ADAR1p110 to JAK2/STAT-inducible ADAR1p150 RNA editase activation contributes to MPN progression. Pre-LSC evolution to LSC was characterized by ADAR1p150 upregulation, distinctive RNA editome patterns, STAT3 hyper-editing, increased replating as a measure of self-renewal. Moreover, LSC generation was typified by beta-catenin self-renewal pathway upregulation, which was recapitulated by lentiviral ADAR1p150 overexpression and reversed by lentiviral ADAR1p150 shRNA knockdown. Our studyunderscores the importance of inflammatory-cytokine fueled enzymatic mutagenesis in human MPN pre-LSC evolution to LSC. Thus, this study sets the stage for developing predictive RNA editome biomarkers of LSC generation to guidetherapeutic strategies aimed at preventing progression of hematopoietic malignancies.

Disclosures

Crews: Ionis Pharmaceuticals: Research Funding.
Low-Dose Methotrexate Could Reduce Severe Early Immune Reactions but Probably Increase Garft Failure in Umbilical Cord Blood Transplantation
Authors: Huilan Liu , Zhu Xiaoyu , Song Kaidi , Wan Xiang , Sun Guangyu , Tang Baolin , Tong Juan , Yao Wen , Zhang Xuhan , Geng Liangquan , Sun Zimin
Published: November 2019

Early immune disorders, including preengraftment syndrome (PES)and acute graft-versus-host disease(aGvHD)are problematic in cord blood transplantation (CBT), and optimal prophylaxis has not been established. Here, we prospectively investigated whether intensive GvHD prophylaxis by adding low-dose methotrexate (MTX)at day 3 after CBThas a prognostic impact on CBT. 16 consecutive single-unit CBT recipients treated for high-risk hematologic malignancies (10 cases) and non-malignancies (6 cases) between February 2019 and March 2019. The patients, 6 female and 10 males, had a median age of 9 years (range 4-40) and a median weight of 26 kg (range 14-68). Myeloablative preparative regimen comprised fludarabine, busulfan and cyclophosphamide for malignancies and reduced-intensity-conditioning comprised fludarabine, cyclophosphamide and total-body irradiation (4 Gy) for non-malignancies. Graft-versus-host disease (GvHD) prophylaxis was cyclosporine and mycophenolate mofetil plus MTX 3 mg/m2 on day+3. 14 patients achieved engraftment at a median of day 19 and had more than 95% (complete) donor chimerism on day+14.Two patients without hematopoietic reconstitution were engrafted after a second CBT. PES was characterized by high-grade fever, rash, pulmonary edema, weight gain, liver and renal dysfunction developed on a median of day 8 in 11 of the 16 evaluable patients, including 2 who did not achieve engraftment. The PES patients received intravenous corticosteroid at a median dose of 1 mg/kg/day, and of these, 2 patients were requiredBasiliximab to improve clinical symptoms. Grade I to IV and aGvHD developed in 4 and only 1 patient developed severe aGvHD with Grade Ⅲ. Follow-up so far, one death occurred at day 103 because of pneumonia. 15 patients are alive well at a median follow-up of 135 days (131-163). Adding low-dose MTX at day +3 may be offer one optimal regimen to reduce severe early immune reactions and improve outcomes in CBT.

Disclosures

No relevant conflicts of interest to declare.
Cost Effective Cardio Protective Intervention in Haploidentical Hematopoietic Stem Cell Transplantation in Children - a Study from a Tertiary Care Centre in India
Authors: Raj Revathi , Uppuluri Ramya , Vellaichamy Venkateswaran , Ravichandran Nikila , Melarkode Ramanan Kesavan
Published: November 2019

Cost effective cardio protective intervention in haploidentical hematopoietic stem cell transplantation in children - A study from a tertiary care center in India

Haploidentical hematopoietic stem cell transplantation with post transplant cyclophosphamide (PTCy) is increasingly being offered to children with life threatening blood disorders with no matched family or unrelated donors. High dose cyclophophamide is an alkylating agent known to cause severe cardiac toxicity. Hepatic metabolism of cyclophosphamide results in the production of phosphoramide which is toxic to the myocardium and acrolein which is a bladder irritant. The metabolites deplete antioxidants and augment the inflammatory damage to the cardiac myocytes. N-acetylcysteine (NAC) helps replenish the oxidant pool in the body and provides cardio protective and overall superior outcomes.

We present retrospective data on 103 children between the ages of nine months to eighteen years who underwent haploidentical transplantation with PTCy between May 2015 and May 2019. All children received 50 mg/kg of cyclophosphamide on day 3 and day 4 after infusion of stem cells with Mesna starting with the first dose of cyclophosphamide as a continuous infusion till 24 hours after the second cyclophosphamide dose. All children had a cardiac ECHO performed by a pediatric cardiologist before conditioning and on day 5 after completion of Mesna. The first 30 children (Group 1) did not receive continuous infusion of NAC and the subsequent 83 children (Group 2) were treated with NAC at a dose of 10 mg/kg/ hour from the start of cyclophosphamide. Disease free survival, graft rejection and cardio toxicity were recorded. The follow up period ranged from 3 months to 4 years. In Group 1 the disease free survival was 46%, graft rejection rate was 13.5% and adverse cardiac events occurred in 10%. In Group 2 the disease fee survival was 67.5%, graft rejection rate was 3.6% and there were no adverse cardiac events.

In conclusion, the introduction of N-acetylcysteine provides a simple and yet cost effective way to eliminate cardiac toxicity even in infants and small children. This is the first interventional study to help safe delivery of a chemotherapeutic agent at a cost of 25 USD for a 10 kg child. We would recommend the use of NAC in all patients undergoing PTCy based transplantation.

Disclosures

No relevant conflicts of interest to declare.
Short-Term Noncryopreserved Hematopoietic Stem Cells: Single Center Experience of Autologous Transplantation
Authors: Voloshin Sergei , Garifullin Andrey , Kuzyaeva Anastasiya , Shuvaev Vasily , Schmidt Alexander , Kuvshinov Alexey , Balashova Valentina , Chubukina Zhanna , Belyakova Elena , Rysev Georgiy , Chechetkin Alexander , Rugal' Viktor
Published: November 2019

Background: High-dose chemotherapy (HDCT) followed by autologous transplantation of hematopoietic stem cells (auto-HSCT) is an important component of treatment in multiple myeloma (MM). There is a standard method of controlled cryopreservation of HSC suspension. We found that the storage of native HSC suspension with temperature fluctuations from +3 °C to +5 °C during 72 - 120 hours does not significantly affect the content of CD34+ cells in the product, the index 7AAD- (7-AAD (7-aminoactinomycin - D) is a fluorescent marker that penetrates damaged cell membranes and binds to double-stranded DNA. Through 7AAD does not penetrate intact membranes, so living cells are not stained 7AAD with flow cytometry), and colony-forming ability (CFA) of HSC, as well as the recovery time of hematopoiesis in MM patients after auto-HSCT.

Aim: To evaluate the effectiveness and safety of the method of storage of non-cryopreserved peripheral blood stem cells.

Methods: 39 patients with MM were included in this study(male/female ratio 1.36:1). All the patients get standard immunochemotherapy programs and were in remission at the time of auto-HSCT. Patients were divided into two groups depending on the method of stem cell storage: group 1 - non-cryopreserved (n=20), group 2 - cryopreserved (standard) (n=19). An effectivity and safety were evaluated in such parameters as the number of CD34+ and 7AAD- cells, CFA after apheresis and before reinfusion of HSC. Also, we evaluated the number of platelets concentrate transfusions, the timing of engraftment of granulocytic and megakaryocytic blood sprouts, the length of hospital stays after auto-HSCT.

Results: The results are presented in the comparison table of the evaluated parameters. Our data showed significantly reduce of episodes febrile neutropenia and cases of enteropathy.

Conclusion: Thus, the proposed method of storage of HSC is not inferior to the traditional method with cryopreservation on such parameters as CD34+, 7AAD-, CFA, the number of platelets concentrate transfusions, terms of hematopoiesis restoration, length of hospital stay after HSCT, the number of complications.

Table.

Disclosures

Shuvaev: Fusion Pharma: Consultancy; BMS: Consultancy; Novartis: Consultancy; Pfize: Honoraria.
Brain Anomalies in Children with Severe Factor VIII Deficiency- a Pilot Study
Authors: Al-Huniti Ahmad , Harshman Lyndsay A , Novak Marci , Nopoulos Peggy , Staber Janice M
Published: November 2019

Introduction: Bleeding resulting from factor VIII deficiency (hemophilia A) can occur in any tissue including the brain. While significant progress has been made in prevention and treatment of hemophilia-related joint and muscle disease with prophylaxis, there is a paucity of research to understand, treat, or prevent brain disease in hemophilia patients. People with hemophilia are recognized to be at risk for neuropsychiatric and neurocognitive disorders even when treated with factor VIII (FVIII) primary prophylaxis. This is especially problematic given that brain injury is difficult to detect early, and consequences are often delayed. Therefore, our study aims to evaluate brain structure and cognitive function in pediatric patients with severe FVIII deficiency in order to improve outcomes.

Methods: After IRB approval, a single-center pilot study enrolled seven pediatric subjects with severe FVIII deficiency and 23 healthy controls to evaluate brain structure and function. Inclusion criteria included males aged 6 - 16 years. All subjects with severe hemophilia A had a FVIII level of less than 1% completed at the Iowa Hemophilia and Thrombosis Center. Any subject was excluded due to a known diagnosis of traumatic brain injury, brain tumor, major chromosomal anomalies, or intellectual disability. Subjects with a known diagnosis of intracranial hemorrhage or FVIII inhibitor were excluded. Neurocognitive assessments included the Behavioral Regulation Index of Executive Function (BRIEF) which measures executive function. Magnetic resonance imaging sequences included T1, T2, and diffusion tensor imaging. Freesurfer and Brain Research: Analysis of Images, Networks, and Systems (BRAINS2) software were used to preprocess imaging and generate volumetric data.

Results: Mean age for PWH was 10.4 years and 11.8 years for controls (p = 0.24). Both groups had similar height, weight, and socioeconomic status. Despite normal IQ, PWH demonstrate impaired behavior regulation and globally impaired executive composition compared to controls (p = 0.001 and p = 0.038, respectively). Volumetric brain quantification revealed subjects with hemophilia had decreased whole brain volumes compared to controls (p = 0.019). Our analysis suggests that these differences are in discrete regions. Future analysis will investigate if volumetric differences correlate with neurocognitive assessments.

Conclusions: PWH demonstrated abnormal brain structure and function compared to healthy controls. Further evaluation is warranted to understand why and how FVIII deficiency and its treatment alter neurological outcomes.

Disclosures

Staber: UniQure: Honoraria; Genentech: Honoraria; Bayer: Honoraria; Spark: Honoraria; Novo Nordisk: Honoraria.
Identifying Novel Immunodominant Epitopes within Human Parainfluenza Virus-3 Matrix Protein
Authors: Harris Katherine Marie , Keller Michael , Bollard Catherine M. , Horn Sarah , Lazarski Christopher , Sani Gelina
Published: November 2019

Human Parainfluenza Virus-3 (HPIV3) is a common cause of severe respiratory illness in immunocompromised patients and has no approved antiviral therapies. Recent preclinical studies demonstrated T-cell recognition of HPIV3 viral proteins, and a phase I study adoptively transferring virus specific T cells (VSTs) targeting HPIV3 to patients following bone marrow transplantation is underway (NCT03180216). Hence, we sought to identify immunodominant epitopes within the Matrix protein based on the extent of the T-cell responses.

VSTs were generated from the peripheral blood of healthy donors by ex vivo expansion after stimulation with a 15-mer peptide library encompassing HPIV3 Matrix protein, which has been identified as the HPIV3 antigen eliciting the most robust response (Mclaughlin et al., J Cytotherapy 2016). Epitope mapping was performed using IFN-g ELIspot with combinatorial 15-mer peptide pools (Straathof et al., J Immunology 2005). Flow cytometry was used to characterize the products with intracellular cytokine staining.

Initial mapping with the peptide pools showed activity in 16 of the 19 pools. Pools which elicited activity in 5 or more products had mean spots per well (ASC)/1x10^5 cells range 89 to 217, versus 0.6 ASC/1x10^5 in the negative controls. We identified 13 novel immunodominant epitopes (Table) within Matrix protein identified in 9 tested VST products, with 4 clustered epitopes (peptides 82-85) at the C-terminus, which contains a leucine residue critical for regulating virion release (Ding et al, 2017). Five (55%) donors recognized epitopes in this domain. Individual peptides which elicited activity had mean (ASC)/1x10^5 cells range 13 to 224, versus 0.4 ASC/1x10^5 in the negative controls. VST products were polyfunctional releasing both IFN-γ and TNF-α in response to the newly identified epitopes, and preliminary flow cytometry data showed that the majority of these epitope specific responses were HLA Class II restricted.

In conclusion, we have identified several novel epitopes within the HPIV3 Matrix protein. Comprehensive classification of the breadth and stability of HPIV3 epitopes is critical for the development of third-party "off the shelf" virus specific T cell therapeutics. Therefore, these results will facilitate the development of clinical studies using partially-HLA VSTs as an "off the shelf' treatment for the treatment of immune suppressed patients with severe respiratory infection related to HPIV3.

Figure

Disclosures

Bollard: Cellectis: Membership on an entity's Board of Directors or advisory committees; NexImmune: Equity Ownership; Torque: Equity Ownership; Caballetta: Equity Ownership; Mana Therapeutics: Equity Ownership; self: Patents & Royalties: patent on HIV T cells. Lazarski:self: Patents & Royalties: IP related to virus T cells.
Catheter Related Thrombosis (CRT) in Hematologic Malignancy: A Retrospective Study of Venous Thromboembolism Management
Authors: Desai Ruchi , Tandon Ankita , Jaglal Michael
Published: November 2019

Introduction: Central Venous Catheters (CVC) improve quality of life for patients with hematologic malignancy but long-term presence may result in CRT. Optimal treatment of CRT is uncertain. Here, we focus on a retrospective study of CRT in patients with hematologic malignancy to identify clinical features associated with increased risk for recurrent thrombosis.

Methods: Demographic and management information of patients with CRT and hematologic malignancy was extracted from the medical record of a single academic medical center. Data was described using summary statistics. Nonparametric Cox proportional hazard models were utilized for thrombosis recurrence and bleeding occurrence. Unadjusted models were utilized for CRT treatment, type of anticoagulation, platelet count, and other factors.

Results: 85 patients were included for analysis. Summary statistics are noted in Table 1. Median platelet count differed across initial treatment approach [Anticoagulation (AC): 177 (106.5- 662), Anticoagulation and Catheter Removal (AC+): 112 (61.75-304), Catheter Removal only 24 (20.25-35.5), No treatment 30.5 (16.75-10.8), p<0.0001]. For CRT treatment, patients with AC+ have more than a 660 % chance of thrombosis recurrence compared to AC as reference group. This significant difference is also seen in the Kaplan Meir curves for CRT treat (log-rank p=0.0484), Figure 1. Of note, 15 patients treated with AC+ developed recurrent thrombosis, the majority of which (53%, 8/15) occurred within 100 days. Of these patients, 62% (5/8 patients) had at least 2 days delay in starting anticoagulation while no delays in starting anticoagulation were noted in patients who were primarily treated with AC. Of the 8 patients who received no therapy for CRT, 7/8 patients (87%) had noted brachial (peripheral deep).

Discussion: Significant variability in management of CRT in patients with hematologic malignancy is noted. Platelet count at time of thrombosis may influence treatment decision. Our data indicate that treatment with anticoagulation is superior to treatment without anticoagulation. The difference in thrombosis recurrence seen in patients who received AC v. AC+ could potentially be explained by delays in starting AC in the latter group. Interestingly, patients who did not receive any therapy for CRT had no recurrence, which may be explained by the fact that the majority had peripheral deep thrombosis, which are less likely to have deep recurrences. In addition, type of anticoagulation used for treatment was notHowever, our data is limited by its population of single institutional patients and retrospective nature. Prospective, studies are needed to better delineate differences in outcomes.

Disclosures

Jaglal: NOVARTIS: Consultancy.
Prognostic Validation of Diffuse Large B Cell Lymphoma International Prognostic Index for High Grade B Cell Lymphoma with and without MYC and BCL2 or BCL6 Rearrangements: An Institutional Review
Authors: Kanagalingam Gowthami , Vyas Vrinda , Basnet Alina , Brooks Garry , Smith Alexandria
Published: November 2019

Background:

Diffuse large B cell lymphoma (DLBCL) NOS is the most common histologic subtype of non-Hodgkin lymphoma (NHL). Double hit lymphoma is re-classified as high-grade B cell lymphoma, with and without MYC and BCL2 or BCL6 rearrangements (HGBCL) in 2016. The International Prognostic Index (IPI) is the main prognostic tool for DLBCL NOS. We aimed to identify if IPI index scores can accurately predict the prognosis in patients with HGBCL.

Methods:

We observed the outcome of a cohort of patients (pts) with DLBCL NOS and HGBCL diagnosed and treated from 2008-2019 at SUNY Upstate Medical University. Fisher's exact test was used for the 2X2 comparison analysis.

Results:

24 pts (82.8%) had DLBCL NOS and 5 (17.2%) pts had HGBCL. The median age was 58 yrs (13-81 yrs). 65.5% were males and 79.3% were caucasians. 8 pts had bone marrow involvement, 1 pt had brain metastasis and 12 pts had bulky disease. 15 DLBCL NOS pts (68.2%) and 4 HGBCL pts (80%) had low-intermediate IPI (</=3). 7 DLBCL NOS pts (31.8%) and 1 HGBCL pt (20%) had high IPI (>/=4). 2 DLBCL NOS pts (13.33%) vs 1 HGBCL pt (25%) died in the low-intermediate IPI group. None of the DLBCL NOS pts (0%) vs 1 HGBCL pt (100%) died in the high risk group.

Conclusion:

Our study sample size is very small to identify prognostic relation of IPI score to predict the outcome for HGBCL. Our observation raises the hypothesis that patient with HGBCL might have a worse prognosis despite low IPI score which needs to be validated in a larger study population.

Disclosures

No relevant conflicts of interest to declare.
Platelet Dysfunction Detected Using Rotational Thromboelastometry (ROTEM) in Severely Thrombocytopenic Patients with a Bleeding Phenotype
Authors: Choi Philip Young-Ill , Hicks Sarah , Gardiner Elizabeth E. , Crispin Philip , Slade James , D'Rozario James , Parish Christopher , Coupland Lucy
Published: November 2019

Background: Thrombocytopenia occurs reasonably frequently with the underlying causes being numerous and requiring time to delineate. Patients may present with active bleeding or low platelet counts may be detected incidentally. Treatments are varied, refractory cases are not uncommon, and in those who do respond, relapses occur. The clinician's principal concern is to limit the risk of a life-threatening hemorrhage, however, the platelet count is an unreliable predictor of bleeding risk. Adult patients presenting with platelet counts < 20 x 109/L present the greatest safety dilemma for clinicians. Previous studies using ROTEM in adult ITP patients with platelet counts < 60 x 109/L and paediatric cases with platelet counts < 30 x 109/L demonstrated that clot firmness parameters correlated with bleeding score1. Our aim, therefore, was to perform ROTEM analysis on patients with a platelet count <20 x 109/L and correlate with bleeding status.

Methods: Patients referred to the Haematology Department of The Canberra Hospital with thrombocytopenia were consented to the study. Blood was collected into Na-citrate tubes and analysed within 4 hr of collection. Each patient had an EXTEM and FibTEM assay performed on a ROTEM Delta machine. The measurement of fibrinogen levels was commenced part-way through the study. Data was analysed using PRISM software.

Results: Twenty-five blood samples were analysed from 21 patients (17 primary ITP, 2 secondary ITP, 1 AML, 1 sepsis), 7 samples were treatment naive. Platelet count ranged from 0-17 x 109/L (mean 7.1 +/- 5.7) Even in this severely thrombocytopenic cohort, platelet count was a poor predictor of bleeding risk, as was the fibrin clot amplitude measured with FibTEM. In comparison, the calculated value for platelet contribution to clot formation (EXTEM A10 minus FibTEM A10) plotted against the platelet count demonstrated that subjects with a bleeding phenotype had a reduced platelet contribution to clot formation in comparison to their non-bleeding counterparts (Figure).

Conclusions: ROTEM has the capacity to detect platelet dysfunction associated with bleeding in severely thrombocytopenic patients. Further studies are underway to elucidate determinants of platelet dysfunction such as specific anti-glycoprotein antibodies that may inhibit platelet function.

Reference #1: Lindsey A. Greene et al., British Journal of Haematology 166: 592-600 (2014)

Figure

Disclosures

D'Rozario: Alexion: Honoraria, Membership on an entity's Board of Directors or advisory committees.
Viability of Stem Cells without Cryopreservation: How Good and How Long?
Authors: Khine Mie Mie , Thandar Hnin Mya , Gyi Aye Aye
Published: November 2019

Although cryopreservation of stem cells is a standard practice for autologous haematopoietic progenitor cells transplantation to treat haematologic malignancies, it has many limitations including toxicities of cryo-protectant, requirement for cryopreservation facilities as well as trained personnel which causes barriers for transplant programs in resource limited countries. To overcome these limitations, a newly established first transplant centre at the North Okkalapa General and Teaching Hospital of Myanmar has been using ordinary refrigerator at 4 degree Celsius to store autologous peripheral blood stem cells collected by apheresis during the interval between collection and transplantation for patients with multiple myeloma who underwent high dose melphalan therapy and autologous stem cell rescue. During early days of the centre, in addition to microbiological testing, to ensure the quality of stem cells, serial measurement for stem cell count and viability were made on daily basis till the day of transplant and continued testing if adequate samples of aliquots were available for analysis. Viable CD34+ cells were analyzed by BD FACS Canto II flow cytometer using BD stem cell enumeration kits that incorporate 7-Amino Actinomycin D viability dyes. The results of the viable CD34+counts of 13 myeloma patients, 5 males and 8 females, median age- 57 years, who underwent autologous peripheral blood stem cell transplantation during early periods of the transplant center from May 2014 to January, 2019, were retrospectively reviewed for the quality of stem cells from the non-cryopreserved methods. Mean CD34 + cells enumerated at the time of reinfusion were 3.45 x 106 cells/kg (range 1.45 - 4.87 x 106 cells/kg). The mean viability of stem cells on day of collection (day 0) were 96.13% (range 83.33% -100%) which was almost unchanged one day after collection (day 1) at 95.93% (85.70% -99.24%). On second and third day after collection, mean viability of the collected cells although fallen below 90% were reasonably preserved at 88.7% (81.74-95.61) on day 2 and 88.08% (87.07-89.09) on day 3, and 85.37% on day 4. Median neutrophil engraftment time was 11 days (range 9-12) and that for platelet engraftment was 16 days (range 9-44). There were no graft failure or mortality during first 100 days. It was encouraging to observe that even at the refrigerator temperature at 4degree Celsius, without any additives or manipulation, stem cell viability could be maintained above 85% minimum of four days with good transplant outcome. Further studies are needed to investigate maximum storage period which could allow reasonable viability and function of the stem cells.

Disclosures

No relevant conflicts of interest to declare.
Mouse APOBEC1 cytidine deaminase can induce somatic mutations in chromosomal DNA
Authors: Caval Vincent , Jiao Wenjuan , Berry Noémie , Khalfi Pierre , Pitré Emmanuelle , Thiers Valérie , Vartanian Jean-Pierre , Wain-Hobson Simon , Suspène Rodolphe
Published: November 2019

Background

APOBEC1 (A1) enzymes are cytidine deaminases involved in RNA editing. In addition to this activity, a few A1 enzymes have been shown to be active on single stranded DNA. As two human ssDNA cytidine deaminases APOBEC3A (A3A), APOBEC3B (A3B) and related enzymes across the spectrum of placental mammals have been shown to introduce somatic mutations into nuclear DNA of cancer genomes, we explored the mutagenic threat of A1 cytidine deaminases to chromosomal DNA.

Results

Molecular cloning and expression of various A1 enzymes reveal that the cow, pig, dog, rabbit and mouse A1 have an intracellular ssDNA substrate specificity. However, among all the enzymes studied, mouse A1 appears to be singular, being able to introduce somatic mutations into nuclear DNA with a clear 5’TpC editing context, and to deaminate 5-methylcytidine substituted DNA which are characteristic features of the cancer related mammalian A3A and A3B enzymes. However, mouse A1 activity fails to elicit formation of double stranded DNA breaks, suggesting that mouse A1 possess an attenuated nuclear DNA mutator phenotype reminiscent of human A3B.

Conclusions

At an experimental level mouse APOBEC1 is remarkable among 12 mammalian A1 enzymes in that it represents a source of somatic mutations in mouse genome, potentially fueling oncogenesis. While the orderRodentiais bereft of A3A and A3B like enzymes it seems that APOBEC1 may well substitute for it, albeit remaining much less active. This modifies the paradigm that APOBEC3 and AID enzymes are the sole endogenous mutator enzymes giving rise to off-target editing of mammalian genomes.
Community resource referral needs among African American dementia caregivers in an urban community: a qualitative study
Authors: Abramsohn Emily M. , Jerome Jessica , Paradise Kelsey , Kostas Tia , Spacht Wesley Alexandra , Lindau Stacy Tessler
Published: November 2019

Background

African American caregivers of community-residing persons with dementia are mostly unpaid and have high rates of unmet basic and health needs. The National Alzheimer’s Project Act (NAPA) mandates improved coordination of care for persons with dementia and calls for special attention to racial populations at higher risk for Alzheimer’s Disease or related dementias (ADRD) to decrease health disparities. The purpose of this study is to describe the perceptions of African American caregivers of people with dementia about community resources needed to support caregiving as well as their own self-care.

Methods

Using a qualitative study design, in-depth, semi-structured qualitative interviews were conducted with caregivers (N = 13) at an urban geriatric clinic to elicit community resource needs, barriers to and facilitators of resource use and how to optimize clinical referrals to community resources. Caregivers were shown a community resource referral list (“HealtheRx”) developed for people with dementia and were queried to elicit relevance, gaps and insights to inform delivery of this information in the healthcare setting. Data were iteratively coded and analyzed using directed content analysis. Results represent key themes.

Results

Most caregivers were women (n = 10, 77%) and offspring (n = 8, 62%) of the person with dementia. Community resource needs of these caregivers included social, entertainment, personal self-care and hospice services. Main barriers to resource use were the inability to leave the person with dementia unsupervised and the care recipient’s disinterest in participating in their own self-care. Facilitators of resource use included shared caregiving responsibility and learning about resources from trusted sources. To optimize clinical referrals to resources, caregivers wanted specific eligibility criteria and an indicator of dementia care capability.

Conclusions

African American caregivers in this study identified ways in which community resource referrals by clinicians can be improved to meet their caregiving and self-care needs.
Clinical and whole-exome sequencing findings in two siblings from Hani ethnic minority with congenital glycosylation disorders
Authors: Zhang Zhen , Huang Ti-Long , Ma Jing , He Wen-Ji , Gu Huaiyu
Published: November 2019

Background

PMM2-CDG, is the most common N-linked glycosylation disorder and subtype among all CDG syndromes, which are a series of genetic disorders involving the synthesis and attachment of glycoproteins and glycolipid glycans. The mutations of PMM2-CDG might lead to the loss of PMM2, which is responsible for the conversion of mannose 6- phosphate into mannose 1-phosphate. Most patients with PMM2-CDG have central nervous system involvement, abnormal coagulation, and hepatopathy. The neurological symptoms of PMM2-CDG are intellectual disability (ID), cerebellar ataxia, and peripheral neuropathy. Now, over 100 new CDG cases have been reported. However, each type of CDG is very rare, and CDGs are problematic to diagnose. In addition, few CDGs have been reported in the Chinese population.

Case presentation

Here we present a Hani ethnic minority family including two siblings with congenital glycosylation disorders. Whole-exome sequencing revealed compound heterozygous for one novel mutation (c.241–242 del variant) and previously reported mutation (c.395 T > C) in gene of PMM2. Two mutations were found in proband and her sibling by whole-exome sequencing. The mutations were identified in this family by Sanger sequencing and no mutations were detected in the normal control.

Conclusions

This is the first report to describe mutations in two siblings of Hani ethnic minority which is one of five ethnic groups found only in Yunnan with a population of more than 1 million.
The value of miR-155 as a biomarker for the diagnosis and prognosis of lung cancer: a systematic review with meta-analysis
Authors: Shao Chuchu , Yang Fengming , Qin Zhiqiang , Jing Xinming , Shu Yongqian , Shen Hua
Published: November 2019

Background

Recently, a growing number of studies have reported the coorelation between miR-155 and the diagnosis and prognosis of lung cancer, but results of these researches were still controversial due to insufficient sample size. Thus, we carried out the systematic review and meta-analysis to figure out whether miR-155 could be a screening tool in the detection and prognosis of lung cancer.

Methods

A meta-analysis of 13 articles with 19 studies was performed by retrieving the PubMed, Embase and Web of Science. We screened all correlated literaters until December 1st, 2018. For the diagnosis analysis of miR-155 in lung cancer, sensitivity (SEN), specificity (SPE), positive likelihood ratio (PLR), negative likelihood ratio (NLR), diagnostic odds ratio (DOR) and area under the ROC curve (AUC) were pooled to evaluate the accuracy of miRNA-155 in the diagnosis of lung cancer. For the prognosis analysis of miR-155 in lung cancer, the pooled HRs and 95% CIs of miR-155 for overall survival/disease free survival/progression-free survival (OS/DFS/PFS) were calculated. In addition, Subgroup and meta-regression analyses were performed to distinguish the potential sources of heterogeneity between studies.

Results

For the diagnostic analysis of miR-155 in lung cancer, the pooled SEN and SPE were 0.82 (95% CI: 0.72–0.88) and 0.78 (95% CI: 0.71–0.84), respectively. Besides, the pooled PLR was 3.75 (95% CI: 2.76–5.10), NLR was 0.23 (95% CI: 0.15–0.37), DOR was 15.99 (95% CI: 8.11–31.52) and AUC was 0.87 (95% CI: 0.84–0.90), indicating a significant value of miR-155 in the lung cancer detection. For the prognostic analysis of miR-155 in lung cancer, up-regulated miRNA-155 expression was not significantly associated with a poor OS (pooled HR = 1.26, 95% CI: 0.66–2.40) or DFS/PFS (pooled HR = 1.28, 95% CI: 0.82–1.97).

Conclusions

The present meta-analysis demonstrated that miR-155 could be a potential biomarker for the detection of lung cancer but not an effective biomarker for predicting the outcomes of lung cancer. Furthermore, more well-designed researches with larger cohorts were warranted to confirm the value of miR-155 for the diagnosis and prognosis of lung cancer.
MSCs helped reduce scarring in the cornea after fungal infection when combined with anti-fungal treatment
Authors: Zhou Yue , Chen Yuqing , Wang Suiyue , Qin Fangyuan , Wang Liya
Published: November 2019

Background

Fungal Keratitis (FK) is an infective keratopathy with extremely high blindness rate. The damaging effect of this disease is not only the destruction of corneal tissue during fungal infection, but also the cornea scar formed during the healing period after infection control, which can also seriously affect a patient’s vision. The purpose of the study was to observe the effect of umbilical cord mesenchymal stem cells (uMSCs) on corneal scar formation in FK.

Methods

The FK mouse model was made according to a previously reported method. Natamycin eye drops were used for antifungal treatment 24 h after modeling. There are four groups involved in the study, including control group, FK group, vehicleinj FK group and uMSCsinj FK group. Mice in uMSCsinj FK group received repeated subconjunctival injections of uMSCs for 3 times at the 1d, 4d and 7d after FK modeling. At 14d, 21d and 28d after trauma, clinical observation, histological examination, second harmonic generation and molecular assays were performed.

Results

The uMSCs topical administration reduced corneal scar formation area and corneal opacity, accompanying with decreased corneal thickness and inflammatory cell infiltration, following down-regulated fibrotic-related factors α-SMA, TGFβ1, CTGF, and COLI and finally inhibited phosphorylation of TGFβ1/Smad2 signaling pathway during FK corneal fibrosis.

Conclusion

The results confirmed that uMSCs can improve corneal opacity during the scar formation stage of FK, and exert anti-inflammatory and anti-fibrotic effects.
Prevalence and risk factors of intestinal parasitic infestations among preschool children in Sekota town, Waghimra zone, Ethiopia
Authors: Kassaw Mesfin Wudu , Abebe Ayele Mamo , Tlaye Kenean Getaneh , Zemariam Alemu Birara , Abate Biruk Beletew
Published: November 2019

Background

Intestinal parasitic infestations triggered considerable gastrointestinal morbidity, malnutrition, and mortality worldwide. In particular, young children in developing countries affected most. Helminthiasis infestation accounts for 10–20% of prevalence on preschool children worldwide. Unfortunately, small children below 5 years are uniquely susceptible to intestinal parasitic infestations in poor communities. This is because of children’s behavior like playing with soil and putting hand -to- mouth habit. Thus, the aim of this study was to assess the prevalence and risk factors of intestinal parasitic infestations among preschool children in Sekota town, Ethiopia.

Methods

A community-based cross-sectional study was carried out on 378 preschool children in Sekota town from February 15 – March 10/2019. Stool specimens were collected and examined for intestinal parasites using wet mount and formal ether concentration technique. The risk factors of intestinal parasites were assessed using a pretested structured questionnaire. The data were entered and analyzed using Epi-data version 4.2.0.0 and SPSS-version 23 statistical software respectively. Both bivariable and multivariable analysis was carried out, and potential co-linearity was tested for closely similar variables. Variables with P value less than 0.05 in multivariable analysis was considered as statistically significant and reported with 95% CI and odds ratio.

Results

The prevalence of intestinal parasitic infestations in Sekota town on wet mount and formal ether concentration techniques was 83(21.9%), (95% CI, 17.7–26.3%) and 113(29.9%), (95% CI, 25.1–34.8%) respectively.

In multivariable analysis, not taking medication as periodical deworming (AOR, 95% CI), (2.5, 1.5–4.3), presence of animals in the living room (AOR, 95% CI) (3.1, 1.8–5.3), and being a government employee as an occupation (AOR, 95% CI), (3.4, 1.1–10.0) were increasing the odds of intestinal parasitic infestations.

Conclusions

The prevalence of intestinal parasitic infestations in Sekota town is high, which is a public health problem. The risk factors that contributed to intestinal parasitic infestations in this study were preventable and modifiable. Therefore, the concerned bodies need to emphasis on periodical deworming and keeping animals in separate room.