bluuebunny/crossref_metadata_2025 huggingface dataset:
Efficacy of Immunomodulatory Therapy with All-Trans Retinoid Acid for Adults with Chronic Immune Thrombocytopenia
Authors: Dai Lan , Wang Zhaoyue , Zhang Ri , Bai Xia , Zhu Mingqing , Ruan Changgeng
Published: October 2019

Abstract 3335

Objective:

Immune thrombocytopenia (ITP) is an immune-mediated disorder characterized by antibody- and cell-mediated platelet destruction and impaired platelet production. In addition of antibody-mediated platelet clearance, T-cell is also involved in platelet destruction or marrow suppression. During immune responses CD4+ T cells can differentiate into a range of cell types, including T helper type 1 (Th1), Th2, Th17 and regulatory T cells (Treg). The goal of the present study is to investigate the therapeutic effects of all-trans-retinoic acid (ATRA) plus prednisone treatment on adult refractory idiopathic thrombocytopenic purpura (RITP) and to further explore the underlying mechanisms.

Methods:

All 35 patients were treated with ATRA at fixed dose of 10 mg/tid plus prednisone and were started after discontinuation of other previous ITP treatment. Peripheral blood samples were collected from 35 RITP patients and 20 healthy subjects. The concentrations of the peripheral blood CD 4+ T cells (Th1, Th2, TH17, TREG) were analyzed by flow cytometry, the levels of cytokines were confirmed by enzyme-linked immunoassay (ELISA), and the expression of T-bet, GATA-3, RORγt, FOXP3 were detected by semiquantitative reverse-transcription polymerase chain reaction (RT-PCR) in 35 RITP patients before and after treatments, with 20 normal individuals as respective controls.

Results:

The overall response rate of ATRA plus prednisone treatment in RITP patients was 54.3%, 28.6%(n=10) of patients with a complete response and 25.7%(n=9) of patients with a partial response. Among the 19 responders, the mean platelet count was 34+13×106/ml before ATRA therapy and 106+29×106/ml after treatment. No severe adverse effects happened. The levels of regulory T cells were significantly increased in patients after ATRA plus prednisone treatment (P < 0.05); the levels of Th1, Th2, Th17 were not significantly improved (P >0.05) in effective patients after treatments in flow cytometry analysis. The concentrations of IL-10, TGF-β was increased (P<0.05), whereas IFN-γ, IL-4, IL-17 factor showed no obvious change in the effective groups after treatment (P > 0.05). The expression levels of Foxp3 enhanced dramatically after treatment in real-time RT-PCR analysis (P < 0.05). However, the concentrations of T-bet, GATA-3 and ROR-γ showed no obvious change after the therapy in real-time RT-PCR analysis(P > 0.05).

Conclusions:

54.3% of RITP patients recovered after ATRA plus prednisone treatments. The therapeutic effects of ATRA plus prednisone may be involved in the increased levels of regulory T cells in peripheral blood through increased expression of TGF-β, IL-10, and Foxp3. In comparison, the therapeutic effects may not be attributed to the expression of TH1, TH2, TH17, IFN-γ, IL-4, IL-17, T-bet, GATA-3, RORγt.

Disclosures:

No relevant conflicts of interest to declare.
Measurement of the Absolute Immature Platelet Number Reflects Marrow Production and Is Not Impacted by Platelet Transfusion
Authors: Bat Taha , Leitman Susan F. , Calvo Katherine R , Chauvet Donna , Dunbar Cynthia E.
Published: October 2019

Abstract 3425

Background

The ability to distinguish increased platelet destruction from platelet hypo-production is important in the care of patients with bone marrow failure syndromes and patients receiving high dose chemotherapy. The measurement of immature circulating platelets based on RNA content using an automated counter is now feasible. This study evaluated the impact of recent platelet transfusion on measurement of immature platelet parameters.

Study Design and Methods

The immature platelet fraction (IPF) and absolute immature platelet number (AIPN) were measured using the Sysmex XE-5000 analyzer prior to and following platelet transfusion in 9 transfusion-dependent patients with marrow failure secondary to aplastic anemia, myelodysplasia or transplantation conditioning. IPF and AIPN were also measured serially over 5 days of storage in 3 plateletpheresis components collected from normal donors.

Results

Platelet transfusion did not significantly change the mean AIPN in transfused patients. In contrast, IPF decreased significantly from 6.6 ±4.6% at day -1 to 2.3 ±1.4% at day 0 before returning to 4.3 ±2.3% at day +1. In the platelet component, AIPN and IPF% increased significantly over 5 days of storage, most likely due to an artifact of the staining and detection process for stored platelets, no longer detected in vivo once the platelets were transfused.

Conclusion

Platelet transfusion decreases the IPF due to the resultant increase in circulating platelet count. However, platelet transfusion does not change the circulating absolute immature platelet number (AIPN), validating this assay as a reflection of ongoing platelet production by the bone marrow in various clinical settings, regardless of proximity to platelet transfusion.

Disclosures:

No relevant conflicts of interest to declare.
Vitamin K3 Is a Potential Ultraviolet Photosensitizer for Pathogen Reduction of Human Platelets and Plasma
Authors: Xu Fei , Vostal Jaroslav
Published: October 2019

Abstract 3434

Human platelets are stored up to 5 days at room temperature and may support bacterial growth before transfusion. Transfusion of bacterially contaminated platelets remains the highest transfusion transmitted infectious disease risk today. One approach to reducing this risk is the development of safe and effective pathogen reduction methodologies. We evaluated UV light (A and B) with vitamin K3 (VK3) as a photosensitizer for efficacy in reducing bacterial growth in platelet concentrates and plasma.

Six species of bacteria, including Bacillus cereus, Pseudomonas aeruginosa, Staphylococcus aureus, Staphylococcus epidermidis, Klebsiella pneumoniae and Escherichia coli, were spiked respectively into phosphate buffered saline (PBS) buffer and exposed to UVA irradiation before bacterial growth was determined. As shown in Figure 1, the bacterial colony forming units (CFU) were reduced with increasing VK3 concentrations and UVA dosages. The VK3 at 400 uM plus UVA-radiation (2.88 J/cm2) significantly reduced the bacterial CFU in PBS spiked with bacteria.

The same six-bacteria panel were spiked respectively into platelet concentrates (PCs) diluted with platelet additive solution and mixed with 400 uM VK3 before the UVA irradiation. As shown in Fig 2a, the bacteria CFU were reduced with increasing dosage of UVA but efficacy declined with lower dilution of PCs. At VK3 concentration of 400 uM and PC diluted to 20% a UVA-dose of 5.76 J/cm2 significantly reduced bacterial CFUs in PCs spiked with E coli, K pneumoniae, P aeruginosa and S epidermidis respectively. Similar results were obtained with the same six-bacteria panel spiked into plasma as shown in Fig 2b.

UVB and VK3 combination had a biphasic inhibitory effect as shown in Fig 3. The VK3 at 200 uM plus UVB-radiation (0.015 J/cm2) also significantly reduced the CFU from the same six-bacteria panel spiked into PBS. However higher concentrations of 1600 uM VK3 could partially reverse the bacterial growth inhibition under UVB-radiation at 0.0038, 0.0075, and 0.015 J/cm2 for E coli, K pneumoniae and S aureus bacterial organisms.

These findings suggest that VK3 may serve as an effective UVA photosensitizer for pathogen reduction of human platelets. With UVB light, lower concentrations of VK3 are effective at inhibiting bacterial proliferation but higher concentrations of VK3 may serve as a weak UVB blocker. Additional studies will need to be conducted to determine the impact of UV light and VK3 on platelet in vitro and in vivo performance.

“This abstract reflects the views of the author and should not be construed to represent FDA's views or policies.”

Disclosures:

No relevant conflicts of interest to declare.
Overexpression of Ribosomal Proteins Is Insufficient for p53 Pathway Induction in Cell Lines and Primary CD34+ Cells
Authors: Raiser David M. , Ebert Benjamin L.
Published: October 2019

Abstract 3479

Diseases of ribosome dysfunction (ribosomopathies), such as Diamond-Blackfan anemia and 5q- syndrome, are caused by deficiencies in one or more ribosomal proteins (RPs) and are characterized by impaired erythropoiesis. The p53 pathway has been identified as a central player in the ribosomopathy disease phenotypes, but the precise mechanism by which p53 is aberrantly induced in the setting of RP deficiency has not been fully elucidated. The current hypothesis for this induction is that, in the context of RP deficiency, cellular levels of free (non-ribosome bound) RPs rise, and a subset of RPs with MDM2-binding ability interact with and inhibit the negative regulatory activity of MDM2 on p53, resulting in p53 stabilization. RPs that have been implicated in this capacity include rpL5, rpL11, rpL23a, and rpS7. This hypothesis also suggests that overabundance of these free ribosomal proteins might be sufficient to induce p53 activity, even in the absence of ribosome protein deficiency. To test this specific aspect of this hypothesis, we generated lentiviral expression vectors carrying the full-length cDNAs for rpL5, rpL11, rpL23a, and rpS7, as well as ARF (which served as a positive control, as ARF is known to induce p53 by binding and inhibiting MDM2), and we used each virus to individually infect A549 cells. Despite increased levels of transcript and protein observed for each of the cDNAs expressed, only ARF overexpression resulted in p53 stabilization and p21 induction. Similarly, when RPs were overexpressed in each pairwise combination, none of the RP pairs tested induced p53 above baseline levels. We also tested all of the overexpression constructs in primary human cord blood-derived CD34+ hematopoietic stem and progenitor cells (HSPCs), and p53 pathway induction was only observed with overexpression of ARF. These data suggest that overabundance of these specific ribosomal proteins alone is insufficient for p53 pathway induction via MDM2 inhibition.

Disclosures:

Ebert: Celgene: Consultancy; Genoptix: Consultancy.
Deregulated Expression of HDAC9 in B-Cells Leads to Lymphoproliferative Disorders As Well As Germinal Center and Post-Germinal Center Derived Lymphomas
Authors: Gil Veronica Sylvina , Stengel Sven , Bhagat Govind , Howell Louise , Catovsky Daniel , Pasqualucci Laura , Petrie Kevin , Zelent Arthur
Published: October 2019

Abstract 3505

Histone Deacetylase 9 (HDAC9) is a class IIa enzyme that, within the hematopoietic system, is preferentially expressed in the B-cell lineage. Previously we have observed that, compared with normal cellular counterparts, expression of HDAC9 appears deregulated in B-lymphoid tumors (Petrie et al., JBC 278: 16059, 2003 and unpublished results). To examine the role of HDAC9 in B-cell transformation we generated a mouse model that constitutively expresses human HDAC9 from early stages of B-cell development under the control of the immunoglobulin heavy chain (IgH) enhancer (Eμ). We demonstrate that from six months of age (6–12 months), Eμ-HDAC9 transgenic mice displayed splenomegaly in a statistically significant fraction of cases (3/17, 18%), which was associated with histopathologic and immunophenotypic evidence of lymphoproliferative disease (LPD) (p<0.0001). Later in life (12 to 23 months of age), 24% (19/78) of Eμ-HDAC9 mice developed a spectrum of mature clonal B-cell lymphoproliferations including monoclonal B-cell lymphocytosis (MBL, 12/78, 15%), splenic marginal zone lymphoma (SMZL, 4/78, 5%) and diffuse large B-cell lymphoma (DLBCL, 3/78, 4%). In contrast, a low background incidence of these malignancies (2%) was observed in control mice (p = 0.0008). Importantly, the HDAC9 gene is located within chromosomal region 7p21, which is a common target of frequent genomic gains in B-cell non-Hodgkin lymphoma (B-NHL) cases and B-NHL cell lines. Consistently, copy number gains in the 7p21.1 chromosomal region were found in 25/79 (32%) primary DLBCL cases analyzed (including one B-NHL patient with a copy number gain only within the 5' end of the HDAC9 gene). Mechanism underlying the role of HDAC9 in lymphomagenesis appears to involve deacetylation of both p53 (leading to deactivation of p53) and BCL6 (leading to activation of BCL6), consistent with a role of a general hypoacetylated state in B-NHL pathogenesis, which is underscored by recently identified inactivating mutations in a number of key histone acetyltransferases. In summary, aberrant expression of HDAC9 leads to the development of B-cell lymphoproliferations, including germinal center (GC) and post-GC lymphomas, showing similarities with those occurring in humans. These findings have relevant therapeutic implications in view of current attempts to use HDAC inhibitors as anti-lymphoma drugs.

Disclosures:

No relevant conflicts of interest to declare.
Mir-126 and Mir-195-Mediated Control of B Cell Fate in Leukemic and Normal Cells As a Potential Alternative for Transcriptional Factor
Authors: Kotani Ai , Okuyama Kazuki , Chanda Bidisha , Ikawa Tomokatsu , Kawamoto Hiroshi , Ando Kiyoshi , Tojo Arinobu , Lodish Harvey F.
Published: October 2019

Abstract 3533

microRNAs (miRNAs) control many developmental and physiological processes. However, it is assumed to work as a fine tuner in cell fate determination, which has been shown to be regulated by transcription factors.

Here, we challenge this canonical notion.

miR-126 is downregulated in MLL-AF4 ALL, biphenotypic leukemia expressing both B cell and myeloid markers, compared with other types of ALLs. CD19 and CD20, B cell differentiation markers, were upregulated when miR-126 is reexpressed in MLL-AF4 ALLs (Figure 1). Interestingly, we found that miR-126, in leukemic cells, induces B cell differentiation (Figure 2a) without changing the expression of E2A, EBF1, or PAX5, (Figure 2b) which are assumed to be critical regulators of B cell development indicating that miR-126 induced B cell differentiation independently of transcriptional factors, such as PAX5, E2A, and EBF1. To test the hypothesis, we tried to rescue block of B cell differentiation in EBF1 knock-out hematopoietic progenitor cells (EBF1 KO HPCs) by exogenous expression of miR-126. As a result, significant upregulation of B cell markers, such as B220, RAG1/2, and CD79a/b was induced by miR-126 in EBF1 KO HPCs which show complete block at pre-pro B cell stage.(Figure 3a) Moreover, miR-126 increased cell proliferation, indicating EBF1 KO HPCs differentiate into large preB cells. Moreover, miR-195, which is upregulated toward pro B cells then downregulated in B cell differentiation, also can partially rescue block of B cell differentiation in EBF1 KO HPCs. (Figure 3b) The block in B lymphopoiesis imposed by the absence of E2A or Pax5 can be overcome by exogenous expression of EBF1. Conversely E2A or Pax5 failed to compensate the effect of EBF1. Thereafter miR-126 and miR-195 potentiates to play more roles than E2A or Pax5 in this system and act as more than a fine tuner in B cell differentiation. Our results elucidate a novel mechanism for the regulation of cell fate independent of transcriptional factor, establish an important role for miRNAs in mammalian lineage specification, and suggest that miRNA could be a new possible therapeutic target for dedifferentiation induced by deregulation of transcriptional factors in ALL, which calls for further studies.

Disclosures:

No relevant conflicts of interest to declare.
Results of a Prospective Phase II Randomized Study of Deferasirox (Exjade®) to Prevent Iatrogenic Iron Overload in Patients Undertaking Induction/Consolidation Chemotherapy for Acute Myeloid Leukaemia (HREC/10/QRBW/135)
Authors: Kennedy Glen A , Morris Kirk , Pillai Elango , Curley Cameron , Butler Jason P. , Durrant Simon
Published: October 2019

Abstract 3577

Background:

Iron overload, as measured by raised serum ferritin, is recognised as a significant risk factor for excess transplant related mortality (TRM) in allogeneic stem cell transplantation (SCT). Due to blood transfusions received during initial induction/consolidation chemotherapy, almost all patients undertaking SCT with AML have significantly raised ferritin pre-SCT.

Aims:

To determine the efficacy of deferasirox (Exjade®) in preventing iatrogenic iron overload in patients receiving induction/consolidation chemotherapy for AML.

Methods:

Prospective randomised phase II study in AML patients planned to receive induction and/or consolidation chemotherapy. Iron studies and CRP were measured pre, mid and post each chemotherapy cycle. Patients were randomised to receive either therapy with deferasirox (starting at 5–10mg/kg/day PO and increasing to maximum 40mg/kg/day PO as tolerated) vs. no deferasirox therapy. Desferasirox was commenced once ferritin increased to >500μg/L and continued until transfusion independence after completion of all chemotherapy. The primary endpoint was ferritin concentration at completion of chemotherapy. Based on historical controls, to detect a difference of >1000μg/L in ferritin between groups a sample size of 14 per arm was required.

Results:

The study was terminated prematurely after 16 patients were enrolled (deferasirox arm n=10; control arm n=6) due to excess grade 2–4 GIT (frequency 1.26 vs. 0.75 events/chemotherapy cycle for deferasirox vs. control arm respectively; p=0.04) and infectious toxicity (frequency 2.22 vs. 1.38 events/chemotherapy cycle for deferasirox vs. control arm respectively; p=0.05) experienced in the deferasirox arm. Overall desferasirox was poorly tolerated, with mean maximum tolerated dose only 13.2mg/kg/day (range 5–20mg/kg/day) and 4/10 patients unable to continue the drug predominantly due to GIT toxicity. Post-treatment ferritin, % change in ferritin and number of transfusions received were no different between arms. Post-chemotherapy CR rate was lower in the desferasirox arm (60% vs. 100%; p=0.234), and all treatment-related deaths (n=3; all infectious-related) occurred in the desferasirox arm (p=0.25). Median OS was similar in both arms (median 450 vs. 476 days for deferasirox vs. control arm respectively; p=0.72).

Conclusions:

Use of desferasirox to prevent iatrogenic iron overload in AML patients undertaking induction/consolidation is ineffective, poorly tolerated, and associated with excess GIT and infectious toxicity.

Disclosures:

No relevant conflicts of interest to declare.
Influence of Ceacams Expression in Graft-Versus-Host Disease After Allogeneic Transplantation
Authors: Koldehoff Michael , Dreesen Janine D. , Elmaagacli Ahmet H. , Beelen Dietrich W. , Singer Bernhard B.
Published: October 2019

Abstract 4490

Background:

The carcinoembryonic antigen (CEA) family is involved in intercellular binding interactions that affect various normal and pathogenic processes associated with cellular growth and differentiation. In human, the CEA family are subdivided into the CEA-related cell-adhesion molecules (CEACAMs) and the pregnancy-specific glycoproteins (PSGs). Never before the influence of CEACAMs on patients who underwent allogeneic hematopoietic stem cell transplantation (HSCT) was evaluated.

Methods:

Here we analyzed in a retrospective study 33 patients (pts) for CEACAMs expression that underwent allogeneic HSCT for various diseases and analyzed their outcome. CEACAMs expressions were performed by flow cytometry and ELISA in whole blood, serum and urine samples.

Results:

We report the analysis of 19 pts with acute leukemia (58%), 5 pts with chronic leukemia (15%), 4 pts with MDS (12%) and 5 pts with advanced NHL (15%). The median age at transplant was 50.5 (range, 18–69) years. In this cohort 12 pts received grafts from HLA-identical siblings (36%), 16 pts from matched (49%) and 5 pts from mismatched (15%) unrelated donors. Transplantat consisted of unmanipulated peripheral blood stem cells (n=26, 79%) or bone marrow (n=7, 21%). Of all pts, 7 (21%) had relapsed after transplant. Among these pts, 31 (94%) developed acute GVHD (21 pts had an acute GvHD of grade ≥2). There was no significant correlation between CEACAMs expression after transplant between the variant leukemic disorders in the whole blood and the urine samples. Analysis of each CEACAMs for relapse showed no statistically differences. For CEACAM-6, we found a moderate up-regulation in pts with acute GvHD ≥2 versus acute GvHD <2 (p<0.1). In pts with severe acute GVHD (grade >3) comparing all other pts, we found significant induction of CEACAM-1 (118.5 ng/ml vs. 198.3 ng/ml, p<0.05) in urine samples and CEACAM-1 (109.2 vs. 157.5 ng/ml, p<0.04) in serum samples. However, no statistic differences were found in the CEACAM-1 in regards to chronic GVHD.

Conclusions:

These results suggest that pts with high levels of CEACAM-1 confirms a relevant association of the development of acute GVHD and CEACAM profiling could be an early indicator of severe acute GvHD.

Disclosures:

No relevant conflicts of interest to declare.
Incidence and Risk Factors for Hypogammaglobulinemia in Pediatric Allogeneic Stem Cell Transplant (SCT) Patients
Authors: Frangoul Haydar , Min Elliot , Lu Zengqi , Mario Mario , Ho Richard , Brown Valerie , Domm Jennifer
Published: October 2019

Abstract 4492

Objectives:

Impairment of antibody immunity correlates with an increased risk of infections after SCT primarily due to encapsulated bacteria. Patients with hypogammaglobulinemia are more prone to recurrent infections that can potentially be fatal. The incidence of this complication is not well known in children following allogeneic SCT. We sought to evaluate the incidence and risk factors of this complication in pediatric recipients of allogeneic SCT.

Methods:

All patients receiving allogeneic SCT at Vanderbilt Children's Hospital between 1999 and 2011 were included in this analysis. Excluded were patients with primary immune deficiency. IgG levels were measured every 2 weeks following SCT for the first 100 days and then monthly thereafter. Hypogammaglobulinemia was defined as IgG<500 mg/dL. All patients with hypogammaglobulinemia received IVIG replacement therapy. Detailed patient and outcome characteristics were recorded.

Results:

A total of 177 patients with a median age of 9 years were included in this analysis. 137 (77%) had malignant diseases and 108 (61%) received cells from an unrelated source. Bone marrow was the stem cell source in 120 (68%) while 30 (17%) received PBSC and 27 (15%) received cord blood. 137 of 177 (77.4%) patients developed hypogammaglobulinemia at a median of 69 days (range 5–339) after transplant. In multivariable analysis patients who developed hypogammaglobulinemia were significantly younger HR=1.05 (95% CI, 1.02–1.1, p=0.002), have malignant disease: HR=2.73: (95% CI:1.6–4.7,p<0.001), received unrelated donor transplant: HR=2.78 (95% CI: 1.75–5.0, p<0.001) and developed acute GVHD: HR=2.24 (95% CI: 1.50–3.32, p<0.001). Patients who developed chronic GVHD and survived for more than 1 year post-transplant were more likely to still be receiving IVIG replacement for hypogammaglobulinemia, (HR=115.5, 95% CI: 14.58–914.1, p<0.001). All patients who are alive >1 year post transplant and are off immune suppression have normal IgG levels and are not requiring replacement therapy.

Conclusions:

Hypogammaglobulinemia is a common complication in pediatric patients following allogeneic SCT. Younger patients, those with malignant diseases and recipients of unrelated donor SCT are at higher risk. Acute and chronic GVHD increased the risk of developing hypogammaglobulinemia. Close monitoring of IgG level should be considered in those at highest risk.

Disclosures:

No relevant conflicts of interest to declare.
Comparative Single-Center Analysis of Hematopoietic Cell Transplantation From Unrelated Umbilical Cord Blood or Haploidentical Related Donor or Mismatched Unrelated Donor in Patients with Hematologic Malignancies
Authors: Fu Chengcheng , Wang Jing , Depei Wu
Published: October 2019

Abstract 4535

One of the truly revolutionary advances in hematopoietic cell transplantation (HCT) is the increasingly successful use of alternative donors, as only 1/4 of patients who require an allogeneic hematopoietic cell transplant will have a HLA-matched sibling donor. A substantial proportion of the need has been met by HLA-matched volunteer unrelated donors, but an unmet need still exists, particularly for people who need a more immediate source of hematopoietic cells in China. Thereby, three alternative graft sources, umbilical cord blood (UCB), haploidentical (haplo) related donor, and mismatched unrelated donor (MMUD) are available.

We report a retrospective analysis of 95 patients with hematologic malignancies who received umbilical cord blood transplantation (UCBT) as a primary unrelated stem-cell source (n=20), or haploidentical (haplo) related donor (n=46), or mismatched unrelated donor (MMUD) (n=29), compare the days of hematopoietic reconstitution & engraftment, and rate of acute graft-versus-host disease (GVHD), and overall survival (OS). The average days of hematopoietic reconstitution (WBC >1.0*10E9) among UCBT recipients were significantly longer than those among haplo/MMUD recipients,(19.77 in UCBT,13.86 in haplo and 11.97 in MMUD, p<0.001). Whereas the average days of full engraftment (STR >95%) among haplo recipients were longer than those among UCBT/MMUD recipients,(24.85 in haplo, 19.29 in UCBT and 20.54 in MMUD, p=0.028). Multivarite analysis demonstrated no apparent differences in the rate of II-IV aGVHD (45% in UCBT, 41.3% in haplo and 55.2% in MMUD, p=0.498), so as to the rate of III-IV aGVHD (30% in UCBT, 41.3% in haplo and 17.2% in MMUD, p=0.543). Overall survival at 2 years was 75% in UCBT, 63% in haplo and 84% in MUUD (p=0.049), and the TRM was 29.4% in UCBT, 39.4% in haplo and 19.2% in MMUD (p=0.245).

It shows that UCB is associated with decreased II-IV aGVHD, but hematologic recovery is slow. Haplo HCT is characterized by donor availability for transplantation and post transplant adoptive cellular immunotherapy, but may be complicated by a high transplantation related mortality (TRM). A MMUD transplant may also be an option, but GVHD may be of greater concern. These data gives us some suggestion for using donor availability to decide on the best option for each individual patient.

Disclosures:

No relevant conflicts of interest to declare.
Continuous High-Dose Idarubicin and Busulfan As Conditioning Regimen for Chinese Patients with Acute Myeloid Leukemia Undergoing Autologous Stem Cell Transplantation
Authors: Ming Hong , Qian Sixuan , Peng Liu , Hanxin Wu , Hongxia Qiu , Hua Lu , Sujiang Zhang , Wei Xu
Published: October 2019

Abstract 4539

Objective:

To investigate the efficacy and toxicity of continuous high-dose idarubicin and busulfan as conditioning regimen for Chinese patients with acute myeloid leukemia (AML) undergoing autologous stem cell transplantation (ASCT).

Methods:

A total of 27 patients with AML were enrolled in our prospective study, including 25 of first complete remission and 2 of second complete remission. Twelve patients were male, and 15 were female, with ages ranging from 14 to 56 (median, 31) years. All the patients were given induction and consolidation treatments and underwent ASCT receiving a continuous infusion of high-dose idarubicin (20 mg/m2 qd, days -13 to -11) and busulfan (oral busulfan 1 mg/kg q6h or intravenous busulfan 0.8 mg/kg q6h, d-5∼-2) as conditioning regimen.

Results:

All the patients acquired hematopoietic reconstitution. The median number of days to neutrophil (©ƒ0.5×109/l) and platelet (©ƒ20×109/l) recovery was 10 and 12, respectively. After a median follow-up of 23 months from diagnosis, the median overall survival (OS) and the median disease-free survival (DFS) were 19(7∼87) and 15(6∼85) months, respectively. A total of 18 patients (73.4%) are alive (17 in continuous complete remission) and 10 (37.4%) relapsed after transplant. Those who underwent standard dose of idarubicin (12 mg/m2 qd, days1–3) and cytarabine (100∼200 mg/m2 qd, days1–7) as induction treatment showed lower relapse rate and better OS and DFS as compared to other induction regimens without significant difference. Patients with favorable karyotypes and molecular biological types acquired better OS and DFS as compared to those with intermediate karyotypes and molecular biological types, but there was no significant difference. Myelosuppression and infections due to neutropenia was the most frequent adverse effects, severe nonhematologic toxicity was not observed in all patients. The patients in our study acquired similar OS and DFS compared to those with intermediate karyotypes and molecular biological types who underwent allogeneic transplantation (HLA identical sibling donor) in first complete remission, and the side effects were minor.

Conclusion:

Continuous high-dose idarubicin and busulfan as conditioning regimen for Chinese patients with AML undergoing ASCT was effect and well tolerable.

Disclosures:

No relevant conflicts of interest to declare.
Cortactin Expression Is Tightly Connected to B-Cell Chronic Lymphocytic Leukemia Aggressiveness
Authors: Gattazzo Cristina , Martini Veronica , Frezzato Federica , Trimarco Valentina , Ave Elisa , Carraro Samuela , Visentin Andrea , Semenzato Gianpietro , Trentin Livio
Published: October 2019

Abstract 4561

Background.

Cortactin is an ubiquitous actin-binding protein, encoded by EMS1 gene and localized in chromosome 11q13 region. This protein is expressed in nearly all mammalian tissues and mostly localizes to dynamic actin structures. As a consequence, cortactin is involved in regulating several actin-dependent processes, including lamellipodium protrusion, trafficking of the key invadopodia proteases and intracellular transducer downstream to kinase-mediated cell signalling upon phosphorylation by Src and Syk tyrosine kinase families. Cortactin is over-expressed in several tumors, such as esophageal squamous cell carcinoma, head and neck squamous cell carcinoma and gastric carcinoma, and so tied to tumor aggressiveness by the promotion of cell invasion and metastasis. We previously showed that cortactin is over-expressed also in neoplastic B cells of patients with B-Cell Chronic Lymphocytic Leukemia (B-CLL). In the present study we investigated the regulation of cortactin activity by the Src kinase Lyn in neoplastic B-CLL cells and the biological implications of cortactin overexpression in this leukemia.

Methods.

Twenty patients and ten normal controls were enrolled. Informed consent was obtained from all patients according to the Declaration of Helsinki. CD19+ lymphocytes were purified from peripheral blood by negative selection using the RosetteSep cells isolation kit (StemCell Technologies). Cortactin localization was performed by Confocal Microscope analysis at basal condition, in presence of the chemotactic stimulus CXCL12 and Src kinase inhibitor PP2. Lyn kinase and cortactin phosphorylation levels were evaluated by western blotting analysis. Migration assay was performed using 3 μm pore filters (Transwell Permeable Supports) in the presence of CXCL12, Src kinase inhibitor PP2 and after cortactin silencing.

Results.

We found that cortactin localization is regulated by Lyn kinase through its phosphorylation. In CLL cells, cortactin was over-phosphorylated and it did not co-localize with actin, as compared to normal cells. PP2-inhibition of Lyn decreased cortactin phosphorylation and triggered its co-localization with actin. Cortactin over-expression proved to be associated to the increased B-CLL spreading through Lyn activity. In fact, not only cortactin over-expression correlated with leukemic cell increased response to CXCL12 (r = 0.9), but also the inhibition of cortactin activity, through PP2 inhibitor or cortactin silencing, drastically reduced neoplastic cell migration after chemotactic triggering.

Conclusions.

These results suggest that cortactin is involved in aggressiveness and spreading of B-CLL cells and that Lyn-cortactin axis could represent an alternative target for the development of new therapeutic strategies.

Disclosures:

No relevant conflicts of interest to declare.
A Splicing Variant of MDM4 Overexpression Plays an Indicator of p53 Aberrations and Marks a Potential Therapeutic Target in Chronic Lymphocytic Leukemia
Authors: Xu Wei , Liu Ling , Fan Lei , Wang Li , Zhang Run , Xu Ji
Published: October 2019

Abstract 4564

Objective

Human homolog of murine double minute 4 (MDM4) belongs to murine double minute (MDM) family. Splicing variant of MDM4 (S-MDM4) is obtained from the deletion of exon 6, which results in an internal deletion of 68 bp. Murine double minute 2 (MDM2) is an analogy of MDM4 and shares highly similar structure with each other. The MDM4 gene plays a crucial role in regulating p53 activity and has been found to overexpress in chronic lymphocytic leukemia (CLL). The purpose of this study was to investigate the prognostic significance of MDM4, and characterize the role of MDM4 in p53 pathway.

Methods

Full-length MDM4 (FL-MDM4), S-MDM4 and MDM2 mRNA expressions were detected by quantitative reverse transcription-polymerase chain reaction (qRT-PCR) in 140 Chinese patients with CLL. The correlation between those MDM expressions and CLL prognostic marker such as clinical stage, immunoglobulin heavy-chain variable region (IGHV) mutational status, ZAP-70, CD38, and chromosomal abnormalities were analyzed. Furthermore, primary CLL cells were treated in vitro with either fludarabine or Nutlin-3, to explore the interaction between p53 status and those MDMs.

Results

FL-MDM4 and S-MDM4 expressions were significantly increased with the del(17p13) (P=0.037 and P=0.006), p53 mutations (P=0.023 and P<0.001) and p53 aberrations (P=0.024 and P<0.001). The correlation between the level of MDM2 expression and p53 status was not observed (P=0.196, P=0.095 and P=0.092, respectively). High level of S-MDM4 mRNA expression was associated with short treatment free survival (TFS) (P=0.004). FL-MDM4 expression was significantly decreased after 24 h treatment with fludarabine (P=0.001), but increased after Nutlin-3-treated (P=0.008) in primary CLL cells without p53 aberrations. Both S-MDM4 and MDM2 expressions were significantly increased after fludarabine-treated in CLL cells without p53 aberrations (P=0.013 and P=0.030). Besides, MDM2 overexpression also occurred in CLL cells with p53 wild type after Nutlin-3-treated (P=0.018). In the primary CLL cells with p53 aberrations or p53 dysfunction, the levels of FL-MDM4, S-MDM4 and MDM2 expressions were not significantly increased or decreased after fludarabine or Nutlin-3 treatment.

Conclusion

S-MDM4 overexpression is an indicator of p53 aberrations in CLL patients, suggesting those patients have a poor prognosis. FL-MDM4 inhibitory effect on p53 can be removed by MDM2-p53 and saved by Nutlin-3, and S-MDM4 overexpression marks a potential therapeutic target in CLL.

Disclosures:

No relevant conflicts of interest to declare.
6q Deletion Helps in the Discrimination Between Symptomatic Waldenström's Macroglobulinemia and Asymptomatic Forms of IgM Monoclonal Gammopathies
Authors: García-Sanz Ramón , Magalhaes Roberto JP , Ocio Enrique M , Giraldo Pilar , de las Heras Natalia , Aguilera Carmen , Hernández Jose Mariano , de Coca Alfonso Garcia , Puig Noemi , Murillo Ilda , Sebastian Elena , González Marcos , Miguel Jesus F. San , Gutierrez Norma
Published: October 2019

Abstract 4566

Cytogenetic abnormalities are present in virtually all cases of Waldenström's macroglobulinemia (WM). Among them, deletion of the long arm of chromosome 6 (6q-) is the most frequently reported in the literature, but the potential impact on disease evolution has not been completely elucidated. In this study we have analyzed the prevalence of this aberration in 203 patients with B-cell lymphoproliferative disorders (B-LPD) producing a monoclonal IgM component. Detection was evaluated by In Situ Fluorescence Hybridization in CD19+ separated cells and results were correlated with disease characteristics. The frequency of 6q21 deletion varied according to the type of B-LPD: 1/42 (2%) in IgM-MGUS, 5/69 (7%) in asymptomatic WM (aWM), 25/83 (30%) in symptomatic WM (sWM) and 0/9 in other IgM B-LPD. Within MGUS and aWM, patients with deletion of 6q displayed a high frequency of adverse prognostic features and required treatment earlier than patients with normal 6q (37 mo vs. +120 mo, p<0.05) although they finally displayed a similar overall survival. Within the sWM, those patients with del(6q) displayed parameters of high tumor burden, such as high levels of b2M and monoclonal paraprotein and more frequent anemia and hypoalbuminemia. Interestingly, there was a correlation between the presence of 6q deletion and the ISS prognostic index (frequency of 6q- among patients stratified in stages 1, 2 and 3 was 24%, 42% and 67% respectively). However, overall survival analysis did not show any difference between patients with and without 6q-, independently if the overall survival was considered globally or free of specific disease mortality. In summary, in IgM monoclonal gammopathies, 6q deletion is associated with more symptomatic forms, higher probability of treatment requirement and high tumor load, although the presence of this abnormality does not represent necessarily an adverse prognosis for OS.

Disclosures:

Ocio: PharmaMar: Patents & Royalties, Research Funding. San Miguel:Celgene: Honoraria; Millenium: Honoraria; Janssen: Honoraria.
Menorrhagia in Women with Severe Bleeding Disorders
Authors: Halimeh Susan , Davies Joanna , Pollard Debra , Abdul-Kadir Rezan
Published: October 2019

Abstract 4626

The management of menorrhagia presents a challenge in women with severe bleeding disorders. Conservative medical management is the first line treatment and most women with severe bleeding disorder require combination treatment. Surgical intervention may ultimately be offered to women in whom medical management has failed and whom no longer desire fertility. Women with low factor levels are at risk of perioperative bleeding complications and may require haemostatic support. A total of 50 women with severe factor deficiencies (less than 20iu/dL) were included in this study. 46 women were registered at the Haemophilia Centre at the Royal Free Hospital in London. Four cases were also included from the Rhine-Ruhr Haemophilia Centre in Duisburg, Germany. We reviewed the occurrence of menorrhagia and the management options that were offered. In those that required surgical intervention, the incidence of postoperative bleeding complications and the requirement for factor concentration was also reviewed.

The bleeding disorders in these women were 34 (68%) with severe factor XI deficiency, 10 (20%) with severe type 1 and type 3 von Willebrand's disease, 4 (8%) with factor VII deficiency, 2 (4%) had factor V or X deficiencies and one (2%) had a combination of factor VI and VIII deficiency. The ISTH/SSC joint working group bleeding assessment tool was used to assess the severity and frequency of bleeding symptoms among this cohort of women. The bleeding scores ranged from −2 to 30 with a median score of 9.5.

In total, 32 out of 50 (64%) women with severe factor deficiency required medical attention for menorrhagia. Medical treatment included hormonal preparations (combined oral contraceptive pill or levonorgestrel intrauterine device), which was used as a first line treatment in 15 out of 32 (46.8%) women. Haemostatic treatment included antifibrinolytic medication such as tranexamic acid, which was used in combination with hormonal therapy. One women required intranasal DDAVP, von Willebrand factor concentrate and tranexamic acid. Failure to control menstrual bleeding occurred in 14 (43.7%) women and surgical intervention was required. 7 out of 14 (50%) women required hysterectomy and the remaining 7 women underwent endometrial ablation. Prophylaxis with factor concentration to cover surgical intervention was given in 8 out of 14 women (64.2%). The remainder received tranexamic acid for 24–48 hours following surgery. Postoperative bleeding occurred in 7 women that had surgical intervention, despite two women receiving prophylaxis. This study highlights the complexity involved in the management of menorrhagia in women with severe bleeding disorders and the high risk of postoperative bleeding.

Disclosures:

No relevant conflicts of interest to declare.
The Ratio of Treg/Th17 Cells Correlates with the Platelet Number in Different Disease State of Primary Immune Thrombocytopenia
Authors: Ji Lili , Li Feng , Zhan Yanxia , Hua Fanli , Zou Shanhua , Wang Weiguang , Cheng Yunfeng
Published: October 2019

Abstract 4649

Background:

Primary immune thrombocytopenia (ITP) is an autoimmune heterogeneous disorder that is characterized by decreased platelet count. Regulatory T (Treg) cells and T helper type 17 (Th17) cells are two subtypes of CD4+T helper (Th) cells. They play opposite roles in immune tolerance and autoimmune diseases, while they share a common differentiation pathway. The imbalance of Treg/Th17 has been demonstrated in several autoimmune diseases. In this study, we aimed to investigate the ratio of the number of Tregs to the number of Th17 cells in ITP patients and evaluate the clinical implications of the alterations in this ratio.

Methods:

Thirty adult patients with newly diagnosed ITP enrolled in this study. Patients who needed treatment had been clinically followed up for 12 months. The percentages of CD4+CD25hiFoxp3+ Treg cells and CD3+CD4+IL-17-producing Th17 cells in these patients and healthy controls (n=17) were longitudinally analyzed by flow cytometry.

Results:

The percentage of Treg cells in ITP patients was significantly lower than that of healthy controls and the percentage of Th17 cells increased significantly at disease onset. It is suggested that the ratio of Treg/Th17 correlated with the disease activity.

Conclusion:

The ratio of Treg/Th17 might be relevant to the clinical diversity of ITP patients, and this Treg/Th17 ratio might have prognostic role in ITP patients.

Disclosures:

No relevant conflicts of interest to declare.
Thrombopoietin Receptor Agonist (ELTROMBOPAG) for Chronic Immune Thrombocytopenic Purpura (ITP) Treatment: 21 Patients in Only One Center
Authors: Román Teresa Álvarez , Salces Monica Martin , Butta Nora V. , Canales Miguel , Fernandez Ihosvany , de la Rua Ana , Yuste Victor Jimenez
Published: October 2019

Abstract 4658

Introduction

ITP management has changed since thrombopoietin receptor agonists (TRAs) were approved for its treatment. Current TRAs are recommended for adults at risk of bleeding who relapse after splenectomy, as well as for those where splenectomy is contraindicated and where, at least, one other therapy has failed.

The goal of this paper is to present the results of the treatment and follow-up of 21 patients treated with Eltrombopag in our Centre during the last year.

Patients

We present 21 patients diagnosed with chronic ITP according to the American Society of Hematology guidelines. 7 of them were males and 14 females. The median age was 70 years (range from 26 to 81 years). Previous splenectomy was undergone in 3 patients. 10 patients had previously received two or more therapies for ITP. 11 patients received TRAs as second line treatment.

Results

On average, platelet counts at the beginning of the treatment with Eltrombopag were 23.000/μL. All patients started with the same doses (50 mg) except one of them who started with 25 mg. Patients only received concurrent ITP therapy (corticosteroids and intravenous immunoglobulins) when platelet counts were under 20,000/μL or bleeding occurred.

8 patients needed rescue medications throughout the treatment with Eltrombopag (7 of them only once). Eltrombopag was withdrawn in a total number of 8 patients due to different reasons: 4 patients were considered as non-responders (3 of them didn't achieve a certain platelet count to prevent major bleeding and, in the fourth one, reducing or suspending the concurrent ITP therapy was not possible. One of these non-responders received Romiplostin and a good response was achieved). In one patient, the therapy was withdrawn due to liver toxicity and in another one, at patient's request. Finally, Eltrombopag was withdrawn in 2 patients because a sustained platelet counts was held.

Conclusion

TRAs are safe, effective and well-tolerated for those patients who relapse after splenectomy or when splenectomy is contraindicated. In our experience, TRAs could be also useful in other diseases like HCV or HIV associated thrombocytopenia. Eltrombopag and Romiplostin bind in different places to the TPO receptor (TPO-R). This circumstance, together with the different activation pathways, may explain that some non-responders to Eltrombopag could respond to Romiplostin and viceversa.

Disclosures:

No relevant conflicts of interest to declare.
Altered Thymocyte Development in Allogeneic in Utero Hematopoietic Cell Transplantation in the Mouse Model
Authors: Li Yan , Vrecenak Jesse D , Watanabe Miho , Li Haiying , Tsai Jacqueline , Peranteau William , Flake Alan W
Published: October 2019

Abstract 4668

Introduction

In Utero Hematopoietic Cell Transplantation (IUHCT) is a promising therapeutic strategy for congenital hematopoietic disorders. While mixed allogeneic hematopoietic chimerism with associated donor specific tolerance is routinely achieved by a predominant mechanism of central deletion, the critical events of donor and host thymocyte development have not been analyzed. In this study, we utilized the murine model of allogeneic IUHCT and analyzed donor and host thymocyte development.

Methods

Bone marrow (BM) cells (10×106) from Foxp3GFP C57/BL6 (B6, H2kb) mice were injected intravenously into Foxp3GFP Balb/c (H2kd) fetuses at embryonic day 14 (E14). At indicated postnatal time points the thymocytes were analyzed by multi-color flow cytometry.

Results

The results demonstrate that the thymic processing of donor BM-derived thymocytes differs significantly from host thymocytes and from thymocyte development in normal B6 and Balb/c control mice. Though each subpopulation of the host's thymocytes showed comparable levels to the normal untransplanted Balb/c mice, the donor BM-derived thymocytes demonstrated significantly higher proportions of CD4+CD8- and CD4-CD8+ single positive cells, and a dramatically lower proportion of CD4+CD8+ double positive cells compared to their donor-derived counterparts, respectively. These discrepancies increased with each analysis time point up to 12 weeks. Immature single positive cells, including both TCRb-CD4+CD8- and TCRb-CD4-CD8+ cells, were significantly higher in the donor-derived thymocytes than the host, indicating the development of the donor BM-derived thymocytes were impeded at the DN-DP stage in this specific allogeneic IUHCT setting. In addition, we also found higher frequencies and more potent suppressive capacity of regulatory T cells in the donor-derived T cell compartment than in the host T cell compartment. In the reversed allogeneic strain combination, i.e. IUHCT of Foxp3GFP Balb/c BM MNCs into the E14 Foxp3GFP B6 recipients, thymocytes of both origins showed imbalanced intra-thymic development that differed from the normal mouse.

Conclusion

Our data in the current study suggests that immune reconstitution of the donor bone marrow derived thymocytes differs from that of the host cells and that of normal mice in an allogeneic IUHCT system. The mechanism of the altered thymic development in allogeneic IUHCT, and the potential importance of these observations for clinical IUHCT remains to be determined.

Bone marrow (BM) cells (10×106) from Foxp3GFP C57/BL6 (B6, H2kb) mice were injected intravenously into Foxp3GFP Balb/c (H2kd) fetuses at embryonic day 14 (E14). At indicated postnatal time points the thymocytes were analyzed by multicolor flow cytometry (Fig. A), and the expression of TCR¦Â in each individual subpopulation in 4-week old mice was measured (Fig. B).

Disclosures:

No relevant conflicts of interest to declare.