bluuebunny/crossref_metadata_2025 huggingface dataset:
Impact of Hydroxyurea On Peri-Operative Management and Outcomes in Children with Sickle Cell Anemia.
Authors: Hayashi Masanori , Calatroni Agustin , Herzberg Brittany , Thornburg Courtney
Published: October 2019

Abstract 2567

Poster Board II-544

Surgical procedures in children with sickle cell anemia (SCA) can be complicated by vasoocclusive events (VOE) such as acute chest syndrome (ACS) and pain. Peri-operative management requires a multidisciplinary approach to provide appropriate pre-operative intravenous hydration and intra- and post-operative monitoring. Transfusion therapy has been controversial. Our institution previously described a low incidence of complications in children who received serial transfusions over 3-4 weeks prior to surgery. Subsequently, an increasing number of children have been prescribed hydroxyurea (HU) to prevent SCA complications. In general, children on HU at our institution only receive a single top-off transfusion the day prior to surgery if their hemoglobin is less than 10 g/dL. We hypothesized that children in the HU group would have a lower number of serial transfusion compared to the non-HU group and that there would be no difference in complications or days to discharge between the two groups. We conducted a single-institution retrospective cohort study of children with SCA, who were age less than 18 years and underwent at least one surgical procedure at Duke University Medical Center between January 1, 2003 and April 30, 2008. Data were abstracted from electronic and written medical records. Descriptive statistics were used to characterize the cohort. Wilcoxon test was used to compare continuous variables and Pearson test was used to compare categorical variables between the non-HU and HU groups. Fifty-three subjects were included (Table 1). The non-HU group was significantly younger than the HU group, but children in the non-HU group were significantly more likely to be transfused pre-operatively, primarily with serial transfusions or erythrocytopheresis, compared to the HU group. One subject in the non-HU group developed a pre-operative delayed hyperhemolytic transfusion reaction. Post-operative complications are detailed in Table 1; the overall rate was low. Two subjects in the HU group developed acute chest syndrome despite pre-operative transfusion; one episode was likely related to underlying asthma and poor response to hydroxyurea; the second was likely related to pain and hypoventilation after laparoscopic splenectomy and tonsillectomy/adenoidectomy. Overall, there were no significant differences in complications and no significant difference in days to discharge between the two groups. In summary, children with SCA on HU may safely undergo surgery without significantly reducing their percent HbS. Nonetheless, attention should still be made towards multidisciplinary effort to reduce intra- and post-operative complications, and clinicians should consider response to HU, pulmonary status and type of surgery when planning peri-operative management in children with SCA on HU.

Disclosures:

Off Label Use: hydroxyurea in young children.
Role of the MAPK and PI3-Kinase/Akt Pathways in the Pre-B Cell Receptor (pre-BCR)-Induced NF-κb Activation and Rag1 and Rag2 Down Regulation.
Authors: Lassoued Kaiss , Fuentes Vincent , Gamlouch Hussein , Bissac Eliane , Marolleau Jean-Pierre , Rochette Jacques , Bouhlal Hicham
Published: October 2019

Abstract 2667

Poster Board II-643

The pre-BCR acts as a critical checkpoint in pre-B cell development and might be also involved in leukemogenesis. Using the 697 and Nalm6 human pre-B cell lines, we have previously shown that pre-BCR stimulation resulted in cell cycle progression associated with activation of number of adaptors and signaling pathways including the PI3-Kinase/Akt, Ras/MAPK, AP1 and the canonical NFkB pathway. We have also demonstrated that Src kinases together with Syk played a crucial role in controlling the pre-BCR-associated functions, acting upstream the above-mentioned signaling pathways. Pre-BCR crosslinking also induced down regulation of Rag1 and Rag2 transcription. In this study we aimed to evaluate the role of MAPK and Akt in the pre-BCR-induced NF-kB activation and Rag1/2 down modulation. For this purpose the 697 pre-B cells and normal bone marrow primary pre-B cells were treated with the U0126 and LY294002, and with MEK1/2 and Akt inhibitors, respectively. A dominant negative form of Akt fused to the HIV1 Tat peptide was also used to inhibit the PI3-Kinase/Akt pathway. We bring evidence that LY294002 could alter the pre-BCR-induced NF-kB activation by inhibiting : i) p105 degradation, ii) p50 NF-kB1 nuclear translocation and, iii) the binding of p50 to an oligonucleotide containing a specific consensus sequence. On the contrary, U0126 significantly enhanced p105 degradation, indicating that MAPK and Akt exerted antagonistic effects on the pre-BCR-induced NF-kB activation. Strikingly the baseline levels of Rag1 and Rag2 transcripts were increased in the LY294002 but not the U0126-treated pre-B cells. Futhermore, both inhibitors were shown to induce a strong increase in the expression of Rag1 and Rag2 transcripts upon pre-BCR crosslinking, suggesting that this receptor exerts dual effects on Rag1/2 expression with a predominant negative regulatory component mediated by both PI3-K and MAPK. No changes in the levels of Pax5, E2A, EBF, IFR4, IRF8, FOXO1, FOXO3, Myb, MAZ, LEF1 and SP1 (transcription factors implied in the regulation of Rag1 and Rag2 transcription) were observed in the pre-BCR stimulated or unstimulated-697 cells, treated or not with the MAPK and Akt inhibitors. Our results suggest that the pre-BCR signaling is a complex and tightly self-controlled process, which deregulation might alter cell growth and survival pathways via NF-kB as well as genomic stability trough Rag1/2 expression.

Disclosures:

No relevant conflicts of interest to declare.
Platelet Derived CD154 Induced by Tat Contributes to HIV-1 Associated Autoimmune Thrombocytopenia.
Authors: Wang Jianhui , Zhang Wei , Li Zongdong
Published: October 2019

Abstract 2669

Poster Board II-645

Enhanced platelet activation was reported in HIV-1 infected patients and strongly correlated with plasma viral load. The HIV-1 Tat protein is able to serve as a ligand of the integrin αvβ3 and several chemokine receptors (CCR2 and CCR3) and to induce downstream signaling in cells express those receptors but the effect of Tat on platelet activation has not been fully investigated yet. Activated platelets are known to express and release a variety of proteins that can modulate the immune system, and platelet derived CD154 is reportedly involved in the development of autoimmune thrombocytopenia. However, the full mechanism underlying HIV-1 induced platelet activation and the biological consequences are not completely understood. In this study, we demonstrate that Tat is able to interact with platelets and β3 integrin by S35 label Tat-platelet binding assay and GST-Tat protein pull-down. We then show that Tat is able to induce platelet activation, up-regulates both CD62P and CD154 in both mouse and human platelets, and results in micro-particle release (as demonstrated by electron microscopy). Tat induces greatly diminished activation in integrin β3 knock out platelets or in platelets pretreated with CCR3 or calcium flux inhibitors, suggesting the requirement of chemokine receptor CCR3, integrin β3 and calcium flux for Tat induced platelet activation. The effect of Tat induced platelet activation on the immune response was studied both in vitro and in vivo. An enhanced immunoglobulin class switch was found in mouse spleen B cells co-cultured with platelets treated with Tat in vitro. In addition, an early antibody response against adenovirus was found in Tat injected mouse immunized with adenovirus suggesting an enhanced immune response in vivo. Thus, we have described a new mechanism in which Tat is able to induce platelet activation and have generated a model in which platelet activation can contribute to the development of HIV-1 associated thrombocytopenia.

Disclosures:

No relevant conflicts of interest to declare.
Granzyme B Produced by Human Plasmacytoid Dendritic Cells Suppresses T Cell Expansion.
Authors: Fabricius Dorit , Vollmer Angelika , Blackwell Sue , Maier Julia , Sontheimer Kai , Beyer Thamara , Mandel Birgit , Lunov Oleg , Tron Kyrylo , Nienhaus G. Ulrich , Simmet Thomas , Debatin Klaus-Michael , Weiner George J , Jahrsdoerfer Bernd
Published: October 2019

Abstract 2674

Poster Board II-650

Human plasmacytoid dendritic cells (pDC) play a central role in regulating adaptive T cell responses in the course of neoplastic, viral and autoimmune disorders. In several of these diseases, elevated extracellular levels of the serine protease granzyme B (GrB) are observed. We found that human pDC can be an abundant source of GrB based on FACS analysis, ELISpot, ELISA, Sensizyme, Western immunoblotting, RT-PCR, and fluorescence microscopy. GrB is actively secreted by pDCs and reaches maximal levels up to two logs higher than those produced by classical GrB producers such as CTL or NK cells. However, pDC GrB production is not accompanied by perforin secretion. Spinning disk confocal microscopy revealed that GrB+ pDC bind to and transfer active GrB to T cells. Importantly, this GrB transfer induces a suppression of T cell proliferation in a GrB-dependent, perforin-independent manner, a process reminiscent of regulatory T cells. GrB expression in pDC is regulated on a transcriptional level by JAK1, STAT3 and STAT5. IL-3 and IL-10 enhance GrB production by pDCs while GrB production is inhibited by toll-like-receptor agonists and CD40 ligand. These findings suggest that GrB production by pDCs is involved in the complex interactions between pDC and T cells and that GrB-secreting pDC may play a regulatory role related to anti-tumor immunity, anti-viral immune responses, and autoimmune processes.

Disclosures:

No relevant conflicts of interest to declare.
Tumor-Derived Microvesicles From Sera of Patients with Acute Myeloid Leukemia Suppresses NK Cell Cytotoxicity Via Transforming Growth Factor Beta-1.
Authors: Szczepanski Miroslaw J , Szajnik Marta E , Czystowska Malgorzata , Mandapathil Magis , Welsh Ann , Whiteside Theresa L , Boyiadzis Michael
Published: October 2019

Abstract 2678

Poster Board II-654

Natural killer (NK) cell cytotoxicity in patients with acute myeloid leukemia (AML) is significantly decreased relative to that in normal controls (NC). However, the mechanisms responsible for low NK cell activity in AML are not known. We considered the possibility that tumor-cell-derived microvesicles (MV) mediate suppression of NK cells. MV originate from the endosomal compartment of activated normal and neoplastic cells. Evidence suggests that tumor-derived MV exert detrimental effects on cells of the immune system and may play a role in tumor progression. To determine their contribution to immune suppression in AML, MV were isolated from sera of patients newly diagnosed with AML prior to any treatment and used to evaluate MV-mediated NK cell suppression. The protein content of MV isolated using exclusion chromatography and ultracentrifugation from sera of 19 AML patients was significantly higher than that of MV isolated from sera of 25 NC (75μg±12/mL vs 1.2μg±0.4/mL, p<0.001 ). MV from AML patients were positive for membrane-associated TGFb-1 and FasL in Western blots, whereas no TGFb-1 or FasL was detected in MV from NC. For functional assays, NK cells sorted from peripheral blood of NC were cultured with MV isolated from sera of the AML patients. A significant decrease in NK cell cytotoxicity was observed after co-incubation with MV (2412 LU before vs 1640 LU after, p<0.002). Concomitantly, a decrease in the expression of the NK cell activating receptor, NKG2D, was observed (57% before vs 38% after, p<0.001). The addition of TGFb1-neutralizing antibody abrogated the effects of MV on the NK cell cytotoxicity and receptor expression. The increased levels in sera of AML patients of MV mediating potent NK cell suppression is likely to compromise anti-tumor immune responses. Therefore, modulation of the levels and functions of MV might provide new immunotherapeutic approaches in AML.

Disclosures:

No relevant conflicts of interest to declare.
Comprehensive Geriatric Assessment-Adapted Chemotherapy in Elderly Patients (>70 years) with Diffuse Large B-Cell Non-Hodgkin's Lymphoma (DLBCL): Final Results and Long Term Follow-up.
Authors: Tirelli Umberto , Fratino Lucia , Balzarotti Monica , Uziel Lilj , Giacalone Annalisa , Ferreri Andres , Santoro Armando , Spina Michele
Published: October 2019

Abstract 2684

Poster Board II-660

Background:

R-CHOP is the standard treatment for elderly patients (pts) with DLBCL. Many pts aged 70 years (yrs) or more are unable to receive R-CHOP and the majority of them are excluded from clinical trials. Comprehensive geriatric assessment (CGA) is an useful instrument to predict the clinical outcome of elderly pts with cancer. Within the GOL (Gruppo Oncoematologico Linfomi) we started a phase II study aiming to evaluate feasibility and activity of a CGA-driven chemotherapy for elderly pts with DLBCL

Material and methods:

Pts with no comorbidity received CHOP/R-CHOP; pts with mild cardiopathy received epirubicin instead of doxorubicin; in pts with moderate/severe cardiopathy the use of anthracyclines was omitted; pts with diabetes did not receive prednisone; in pts with neuropathy vincristine was omitted. The dosage of chemotherapy was decided according to CGA: pts with a good score (ADL=6 and IADL>6) received full doses of CT; pts with an intermediate score (ADL=5 and IADL>4) received 75% of the dose; pts with a poor score (ADL<5 and IADL<5) received 50% of the dose.

Results:

One hundred pts (41 males and 59 females) have been treated. The median age was 75 yrs and stages III-IV were diagnosed in 51% of pts. 61% of pts received full doses of CT; 25% received 75% of dose and 14% received 50% reduced dose; 86% of pts received an anthracycline and 54% rituximab. Toxicity was quite acceptable. Grade 3–4 neutropenia was observed in 30% of pts, mucositis in 12%, and peripheral neuropathy in 9%. Four toxic deaths were observed. Overall, 81% of pts achieved complete remission; with a median follow-up of 50 months, 20% of them have relapsed. The 5 yr-OS, DFS, EFS are 58%, 78% and 50%. It is remarkable that the 5-year specific survival is 72%.

Conclusions:

Our results demonstrate that a CGA-driven approach is feasible in elderly pts with DLBCL. This strategy allows to offer a curative approach to all pts with aggressive NHL, avoiding to under treat pts with a potentially cured disease or over treat pts with severe comorbidities.

Disclosures:

No relevant conflicts of interest to declare.
Phase I/II Study of Escalating Doses of Idarubicin in Combination with Methotrexate, Vindesine and Prednisolone in Patients with Primary Central Nervous System Lymphoma: Results of the GOELAMS LCP99 Multicenter Trial.
Authors: Olivier Gaelle , Gardembas Martine , Delwail Vincent , Le Mevel Annie , Escoffre-Barbe Martine , Brion Annie , Boulet Jean Michel , Solal-Celigny Philippe , Ingrand Pierre , Colombat Philippe
Published: October 2019

Abstract 2694

Poster Board II-670

Introduction:

This study was performed to determine the maximum-tolerated dose (MTD) and the recommended dose (RD) of idarubicin in combination with methotrexate (MTX), vindesine (VDS) and prednisolone (PRED) given to previously untreated patients (pts) (60–70 years-old) with primary central nervous system lymphoma (PCNSL).

Patients and Methods:

MTX and VDS were respectively given at the fixed dose of 3 g/m2 (6-hour intravenous [IV]) and 3 mg/m2 IV both on day 1. PRED was given at the fixed dose of 60 mg/m2 (IV or per os) on day 1 to 5. Idarubicin was administrated IV on day 1 with doses starting at 12 mg/m2 and escalated by 2 mg/m2 increments until MTD. Treatment was repeated every 3 weeks, three times. G-CSF was used. Intrathecal chemotherapy (MTX 15 mg, cytarabine 40 mg ) was performed on day 1. Subsequent whole brain irradiation (30 Gy + 10 Gy on residual mass) was performed. Limiting toxicity was defined as grade 4 neutropenia for more than 7 days or thrombopenia grade 4 or extra haematological toxicity more than grade 2.

Results:

Between 2000 and 2005, 35 immunocompetent pts (19 men, 16 women) with a new diagnosis of PCNSL received 87 cycles at four different idarubicin dose levels. At dose level 1 (12 mg/m2) and level 2 (14 mg/m2), only one of six developed a limiting toxicity. About 3 cases, no limiting toxicity was observed at level 3 (16 mg/m2). Three of six developed a limiting toxicity at level 4 (18 mg/m2) which was considered as MTD. Fourteen patients were enrolled at level 3 to confirm the data. At level 3, main haematological toxicities were neutropenia grade 3 (9%) or grade 4 (40%) and main extra haematological toxicities were infectious disease (5 % grade 3 or 4) and renal failure (grade 2: 9%). Median survival time was 385 days (IC95% 136–679).Overall response rate after radiotherapy was 65.7 % (15 CR, 7 PR).

Conclusion:

Our study suggests that the RD of idarubicin administrated with MTX, VDS and PRED, should be 16 mg/m2 on day 1 every 21-day cycle.

Disclosures:

Delwail: Innate Pharma: Honoraria.
CD23 Receptor Positivity Has No Prognostic Implication in Chronic Lymphocytic Leukemia.
Authors: Aghalar Jahan , Chu Charles , Damle Rajendra N , Tsao Che-Kai , Kohn Nina , Kaufman Matthew
Published: October 2019

Abstract 4397

BACKGROUND

Chronic Lymphocytic Leukemia (CLL) phenotypically expresses CD23, although the percentage of positive cells measured by flow cytometry is variable. We sought to analyze whether the percent of CD23 positive cells in the CLL clone correlates with time to treat (TTT), overall survival (OS) and prognostic markers CD38, ZAP-70, and IGHV mutation status.

METHODS

We retrospectively analyzed the flow cytometry data of 332 CLL patients on the gated population of cells that were CD5 and CD19 positive. Percentage positivity for CD23, CD38, and ZAP-70 was noted. CD38 and ZAP-70 were considered positive at cut-offs of >= 30% and >=20%, respectively. CD23 was considered negative at <30% and positive at >= 30%. IGHV sequence was determined from cDNA and then compared to germline to assess mutation status using IMGT/V-QUEST.

The distributions of time from diagnosis until start of treatment and overall survival were stratified by CD23 positivity, estimated using the product limit method, and compared using the log rank test. Those who had expired without treatment or were alive and not treated at this time point were censored in the TTT analysis. Those who were still alive were censored in the OS analysis. Associations of CD23 positivity with IGHV mutation status, ZAP-70, and CD38 positivity were examined using the chi-square test.

RESULTS

Out of 332 patients, 25 had diminished CD23 expression (<30%) whereas 307 had normal CD23 expression (>30%). There was no difference in time until start of treatment or overall survival based on CD23 %positivity. CD23 %positivity showed no associations with IGHV mutation status, ZAP-70 or CD38 positivity.

CONCLUSION

CD23 percent positivity has no prognostic significance in CLL. There is no correlation between CD23 percent positivity and poor prognostic markers such as CD38, ZAP-70, or IGHV mutation status.

Disclosures:

No relevant conflicts of interest to declare.
Deacetylase Inhibitor Cocktails Provide Striking Synergistic Interactions in Human Leukemia Cells.
Authors: Cea Michele , Cagnetta Antonia , Fruscione Floriana , Bruzzone Santina , Zoppoli Gabriele , Garuti Anna , Moran Eva , Grillo Valentina , Ferrando Fabio , Casciaro Salvatore , Pierri Ivana , Gobbi Marco , Ballestrero Alberto , Patrone Franco , Nencioni Alessio
Published: October 2019

Abstract 4404

Cancer cells almost invariably exhibit aberrant histone deacetylase (HDAC) activity leading to changes in chromatine structure, altered gene expression, poor differentiation, impaired apoptosis and increased proliferation. Accordingly, virtually all the HDAC inhibitors currently available show some degree of antitumor activity in preclinical cancer models and several of these compounds are currently under investigation or already approved for the treatment of human malignancies. Such is the case of the hydroxamic acid derivative suberoylanilide hydroxamic acid (Vorinostat, Zolinza), approved for the treatment of cutaneous T cell lymphomas. Sirtuins are a large family of deacetylases characterized by a unique, NAD+-dependent enzymatic mechanism. In addition to their established role in metabolism and longevity, recent evidence points to an emerging role for sirtuins in carcinogenesis. In the attempt to identify drug combinations that would increase the activity of traditional HDAC inhibitors we have explored the combination of valproic acid (VA) and butyrate (BU) with the sirtuin inhibitors cambinol and sirtinol in primary B-cell chronic lymphocytic leukemia (B-CLL) cells (n=35), acute myelogenous leukemia (AML) cells (n=10) and leukemia cell lines. Cell viability was assessed by propidium iodide staining and flow cytometry. Combination indices were determined using the median-effect method. In leukemia cells, exposure to sirtuin inhibitors synergistically increased VA and BU mediated cytotoxicity. Conversely, these drugs were poorly active and failed to show any cooperation in healthy cells, including peripheral blood mononuclear cells and fibroblasts, suggesting a cancer-specific mode of action. Similar results were obtained by combining VA or BU with the Nampt inhibitor APO866, which reduces intracellular NAD+ levels and thereby prevents sirtuin activity. Remarkably, SIRT1 and SIRT6 inhibition per se did not seem to account for cell demise upon HDAC inhibition since expression of a dominant negative SIRT1 isoform or RNA interference-mediated SIRT6 silencing failed to increase VA and BU activity. Our data indicate a specific requirement by leukemia cells for sustained sirtuin activity when classical HDACs are inhibited. This feature is suitable to be therapeutically exploited by combining sirtuin inhibitors or APO866 with classical HDAC inhibitors especially for the treatment of hematological malignancies.

Disclosures:

No relevant conflicts of interest to declare.
Gene Profiling Mechanisms of Cell Density-Dependent Growth in Myeloid and Lymphoid Leukemia Cell Lines.
Authors: Murohashi Ikuo , Ihara Noriko
Published: October 2019

Abstract 4420

Normal hematopoietic stem cells have been shown to be maintained through interaction with their environmental niches, such as osteoblastic and endothelial ones. The growth of leukemic cells has been shown to be stimulated by environmental niches (paracrine growth) or by cell-to-cell interaction or excreted factors of leukemic cells (autocrine growth). The growth of myeloid (MO7-E and HL-60) and lymphoid (Raji, U-266, Daudi and RPMI-1788) leukemia cell lines cultured at various cell densities in serum free medium (Sigma H 4281) with 1% BSA was evaluated. The cells cultured at higher cell densities (cultured cell densities ≥a 105/ml) showed logarithmic linear increases in cell number, whereas those at lower cell densities (cultured cell densities □… 104/ml) ceased increasing cell number. Supernatants of myeloid leukemia cells stimulated the growth of autologous clonogenic cells, but not those of lymphoid leukemia cells. Neutralizing antibodies (Abs) against various hematopoietic growth factors failed to inhibit cell growth except for anti-VEGF, which significantly decreased HL-60 leukemia cell growth. To clarify the nature of the cultured cell density on the growth of leukemia cells, leukemia cells were cultured at higher cell density (group H, cultured cell densities of 106/ml) or at lower cell density (group L, cultured cell densities 104/ml). After culture of 3-, 6-, 10-, and 24-hr, cells were serially harvested and total cellular RNA was extracted. Gene transcript levels were determined by using Real-Time PCR. Gene transcripts examined in the present study were as follows: polycomb (Bmi1), Hox (HOXA7, HOXA9, HOXB2, HOXB4, Meis 1), Caudal-related (CDX2, 4), Mef2c, c-Myb, Wnt (Wnt 3a, Wnt 5a, β-Catenin, β-Catenin, N-Cadherin), Notch (Notch-1, -2, -3 and Jagged-1, -2), CKI (p14, p15, p16, p18, p21, p27, p57), growth factor (VEGF, IGF-1, -2, Ang-1, -2, SDF-1), growth factor receptor (Flt-1, KDR, neurophilin-1, IGF-1R, Tie-1, -2, CXCR4), and growth related (c-Myc, CyclinD1, Foxo3a) genes. p18 and p21 gene expression was higher in group L compared with group H in two and all five groups, respectively. In contrast, p14 gene expression was higher in group H compared with group L. Any of the p15, p16, p27 and p57 genes was deleted. VEGF gene expression levels at 1-3- hr culture were higher in group H compared with group L. HOX, Meis 1 and Mef2c gene expression levels at 1- to 10- hr culture were higher in group H compared with group L. At 24-hr cultures, transcripts of myeloid and lymphoid cell lines for Bmi-1, Wnt-3a, and β-Catenin were higher, and those of lymphoid cell lines for Notch 1, 2, and 3 were higher in group H compared with group L. Taken together, our present results favor the conclusions that genes related to growth factors and transcription factors are sequentially and differentially expressed through cell-to-cell interaction and excreted autocrine growth factors of leukemia cells.

Disclosures:

No relevant conflicts of interest to declare.
Hyper-Methylation DAZAP2 May Suppress Its Expression in Specific Subtypes of Myeloma.
Authors: Hu Wei-Xin , Qu Qiang , Li Jiang , Ren Wei
Published: October 2019

Abstract 4421

Most malignant features of cancer cells are triggered by activated oncogenes and the loss of tumor suppressors due to mutation or epigenetic inactivation. We previously reported that DAZAP2 was down-regulated in newly diagnosed myeloma (MM) and this may influence MM cell growth and survival. This study was undertaken to evaluate the mechanism of down-regulation DAZAP2 and determine the methylation status and loci of its promoter by using bisulphite genomic-sequencing (BGS) method in KM3 myeloma cell line. Two islands with rich GC box in the promoter of DAZAP2 gene were identified and amplified by using bisulfite-sequencing PCR (BSP). Island 1 spans -472 to -247 including 9 CpG sites (CpGs) and island 2 covers -226 to -13 including 23 CpG sites. The ratio of methylated CpGs in two CpG islands was 46.25%. The above sequences were demethylated and inserted into a pGL2-basic vector. COS-7 cells were transfected with recombinant plasmids and the activity of luciferase was evaluated. The results showed that the CpG island 1 had a weakly transcriptional activity, whereas the CpG island 2 had a strong transcriptional activity (2.38 folds compared with the positive control). The other sequence which covered CpG island 1 and 2 showed a remarkable activity (15.1 folds compared with the positive control). These data indicated that CpG island 2 of DAZAP2 gene may be hypermethylated and suppressed its expression in MM cells. We further correlated DAZAP2 expression with normal plasma cells and malignant myeloma cells, as well as the molecular subtypes which the dataset includes 8 genetic subtypes (MY, PR, LB, MS, HP, CD-1, CD-2, and MF) from 351 newly diagnosed myeloma cases (Zhan, 2006). Interestingly, we extended and confirmed our previous discovery that DAZAP2 was significantly down-regulated in MM cells by using a large uniform dataset (P = 0.004). The low expression of DAZAP2 was especially significant in the subgroups of MY, PR, LB, HP, and CD-1. This study warrants further investigation of DAZAP2 and its potential role in myeloma.

Disclosures:

No relevant conflicts of interest to declare.
Change of the Notch1 Expression Level in Lymphoid Malignancies.
Authors: Zheng Haitao , Yang Lijian , Chen Shao-Hua , Zhong Liye , Li Yangqiu
Published: October 2019

Abstract 4425

Notch gene family encodes single pass transmembrane receptors able to transducer intercellular signals involved in cell-fate determination. Notch signaling is the leading causes to the development of malignant tumors. Notch pathway plays a role in haematopoiesis affecting both stem cells and committed progenitors. The expression of Notch1 in thymic stromal cells and in developing thymocytes is consistent with Notch function. Notch1 is turn on before T-cell commitment along the lymphoid development; a variety of model systems indicates that Notch1 signaling is required for the earliest stages of T-cell commitment in the thymus and that, in its absence, cells develop toward the B-lineage. Overexpression of Notch1 gene was found in T-ALL, however, little is known the expression pattern of Notch1 gene in the different lymphoid malignancies. In the present study, the expression level of Notch1 gene was analyzed in patients with lymphoid malignancies. Real-time PCR with SYBR Green I technique was used for detecting Notch1 gene expression level in peripheral blood mononuclear cells of 43 patients with lymphoid malignancies [including 7 cases with T-ALL, 6 cases with T-NHL, 9 cases with B-ALL, 5 cases with B-CLL, 16 cases with B-NHL] and 20 healthy individuals. β2-microglobulin gene (β2M) was used as an endogenous reference. Relative changes in Notch1 gene expression level were used by the 2-ΔCt×100% ×100% method. The expression level of Notch1 in both T-cell lymphoid malignancies group (Median: 0.897%) and B-cell lymphoid malignancies group (Median: 0.726%) was significant higher than those in the healthy control (Median: 0.288%) (P<0.01, P<0.01). Moreover, the expression level of Notch1 was different in the five types of lymphoid malignancies. That is, the highest in the B-CLL group, secondary in the T-NHL group, the lowest in the B-NHL group. And, it is not significant different between the B-NHL and healthy control groups. In conclusions, Overexpression of Notch1 gene was found not only in T-ALL, but also in T-NHL and B-cell leukemia and suggests that the abnormal overexpression pattern of Notch1 might play an important role in the pathogenesis of lymphoid malignancies. Down-regulation of Notch1 might be considered as target therapeutic strategy for lymphocytic malignancies.

Disclosures:

Zheng: The study was supported by grants from National Natural Science Foundation of China (No. 30871091): Research Funding. Yang:The study was supported by grants from National Natural Science Foundation of China (No. 30871091): Research Funding. Chen:The study was supported by grants from National Natural Science Foundation of China (No. 30871091): Research Funding. Zhong:The study was supported by grants from National Natural Science Foundation of China (No. 30871091): Research Funding. Li:The study was supported by grants from National Natural Science Foundation of China (No. 30871091): Research Funding.
Hemophilia in the Peruvian Social Security System: Report of Five Centers.
Authors: Chumpitaz Gloria , Cauvi Fernando , Navarro Juan Ramon , Pedraza Karina
Published: October 2019

Abstract 4446

The Hemophilia Unit of the Hematology Department of the National Hospital Edgardo Rebagliatti Martins- ESSALUD, is one of the most important Hemophilia Centers through the country, that assists patients not only in its jurisdiction but also a great amount referred from other institutions due the complexity of their treatment, like acquired inhibitor disorders as well as orthopedic and mayor cardiovascular surgeries and severe hemorrhages.

The Hemophilia ESSALUD system is compounded of centers in Lima (02), Callao (01) and provinces (05) (Arequipa, Chiclayo, Trujillo, Piura and Cuzco).

The range of patients with new diagnosis is about 5 to 20 patients per year.

The prevalence of Hemophilia in the last 5 years, accounts 83.38% patients of the global amount with coagulation disorders. Of the 331 patients in treatment, 228 (68.8%) have Hemophilia A. Considering the classification of status severity, 12.5% patients of this group belong to mild status hemophilia, 39.47% patients to moderate status, and 41.43% to severe status.

In relation to Hemophilia B, we account 48 (14.5%) patients; 5.2% corresponds to mild status, 23.68% moderate status and 52.63% severe status.

The rest of the patients have other disorders of coagulation such as Von Willebrand disease and rare coagulation factor deficiency (V, VII, XI and XII).

According to the age group distribution for this series, a mayor proportion of 44.68%of patients belong to the group between 16 to 35 years old. The group above 35 years accounts 30%.The age groups of 6 to 15 years old and the group of 1 to 5 year old account 16.48% and 8.51%, respectively.

Finally, it is important to point out that 37.76% of our hemophiliac patients are in the group of moderate illness status and 44.14% in the group of severe illness status. All the patients receive treatment with plasma-derived coagulation factor concentrates and in some cases recombinant therapy. Children receive prophylactic treatment.

Disclosures:

No relevant conflicts of interest to declare.
Retrospective Review of Life-Threatening Bleeds in Paediatric ITP.
Authors: Killen Maire-Clare , Reeves Marie , Grainger John D.
Published: October 2019

Abstract 4453

Immune thrombocytopenia (ITP) is an uncommon disorder of childhood. Most children recover quickly regardless of intervention, but severe bleeding develops in approximately 3%. Intracranial haemorrhage (ICH) is by far the most feared complication, occurring in 0.1-0.5%. Pharmacotherapy is available to transiently increase platelet counts, but introduction of British clinical guidelines in 1992 has resulted in a sharp decline in the number of children receiving such treatment.

The aim of this survey is to establish the current UK incidence of intracranial and other life-threatening haemorrhages in ITP, and whether or not children developing such complications have other risk factors for bleeding.

UK paediatricians were mailed a questionnaire asking to report the number of cases of ITP per year, and any cases of intracranial haemorrhages (ICH) in the previous ten years or other life-threatening haemorrhages in the past five years. Consultants reporting patients were asked to complete a form requesting detailed information on the child and the circumstances surrounding their bleed.

844 annual cases of ITP were reported from 145 centres; 672 of these were acute presentations and 240 were chronic. 24 cases of ICH and 19 cases of life-threatening bleed were identified, giving an annual incidence of 0.3% and 0.5% respectively. Gastrointestinal haemorrhage was the most commonly reported life-threatening bleed, and in six cases, ICH and life-threatening bleeds occurred in the same child.

The incidence of ICH in ITP may be higher than previously reported in the UK; the reason behind this increase remains unknown and support the need for larger international collaborative studies.

Disclosures:

No relevant conflicts of interest to declare.
GRAFT-VS-LEUKEMIA EFFECT in a MOUSE MODEL of CHRONIC MYELOID LEUKEMIA.
Authors: Lamana María L , Oviedo Alberto , Yañez Rosa , Aldea Montserrat , Rubio Antonio , Bueren Juan A.
Published: October 2019

Abstract 4500

Graft-vs-Host Disease (GVHD) is a frequent and severe complication of allogeneic hematopoietic stem cell transplants (allo-HSCT), mediated by donor's T cells reacting against host antigens. However, donor's T lymphocytes also generate the beneficial Graft-vs-Leukemia effect (GVL) by recognizing tumor antigens as non-self, thus contributing to the eradication of residual leukemic cells. With the purpose of studying factors that could affect the GVHD/GVL effects, we generated a chronic myeloid leukemia (CML) mouse model by the transplantation of lin- bone marrow (BM) cells transduced with a retroviral vector carrying the BCR/ABL and tNGFR marker genes (p210-tNGFR RV) into syngeneic, lethally irradiated, B6D2F1 (H2b/d) mice. Transplanted mice developed chronic myeloid leukemia (CML), characterized by hemorrhagic lungs, hepatomegaly, splenomegaly and granulocytosis. Additionally, NGFR+ leukemic cells were detected in the peripheral blood and bone marrow. All mice died on days +18-20 after infusion of the transduced cells. The co-transplantation of B6D2F1 mice with syngeneic p210-tNGFR transduced lin- BM cells together with allogeneic BM cells from C57Bl/6 mice (H2b/b), determined a slower progression of the disease. In this case, transplanted mice died of leukemia on days +28-72 after the co-infusion.

To investigate in this CML mouse model the GVHD/GVL effect mediated by donor allogeneic T-lymphocytes, splenocytes from allogeneic C57Bl/6 mice were additionally infused together with the allogeneic BM cells and the symgeneic p210-tNGFR transduced lin- cells. No CML signs developed, and no NGFR+ cells were detected in mice receiving the allogeneic T cells. In this experimental group, however, all the animals died from acute GVHD on days +13-36 after the co-transplantation, similar to the GVHD control group that received allogeneic bone marrow cells and T-lymphocytes but not the p210-tNGFR transduced lin- cells.

In previous works, we demonstrated that the infusion of adipose tissue-derived mesenchymal stromal cells (Ad-MSCs) prevented GVHD in a haploidentical hematopoietic progenitor cells transplantation mouse model (Yañez et al, Stem Cells 2006). With the present GVHD/GVL mouse model we are now investigating the impact of the infusion of the immunosuppressive adipose tissue-derived mesenchymal stromal cells on the GVHD/GVL effect mediated by allogeneic T cells.

Disclosures:

No relevant conflicts of interest to declare.
The Effect of High-Resolution Donor-Recipient HLA Matching and Mismatching Frequencies On Choosing Donor in Chinese Population for Unrelated Allo-HSCT.
Authors: He Jun , Chen Zi-Xing , Bao Xiaojing , Qiu Qiaocheng , Yuan Xiaoni , Li Yang
Published: October 2019

Abstract 4503

The relative importance of various human leukocyte antigen (HLA) loci and the resolution level at which they are matched has not been fully defined for unrelated donor (URDs) transplantation. Hematopoietic stem cell transplantation (HCT) from volunteer URDs may give a chance of cure for patients with malignant hematological diseases. Although donor-recipient HLA matching is associated with better outcomes, many are not able to identify an HLA-A, -B, -C, -DRB1, DQB1 matched URD and are faced with choosing the closest matching among the available donors. The Chinese Marrow Donor Program (CMDP) has completed a retrospective high-resolution HLA typing on sufficient patient-donor pairs to analyze high resolution matching and mismatches probability at specific loci. These data are critical for selecting the best available partially HLA-matched donor for patients undergoing HLA-mismatched URD HCT. We have performed high-resolution typing for HLA-A,-B,-C,-DRB1,-DQB1 by using SBT, SSOP and SSP techniques on 1092 donors and 931 patients from the data base of CMDP. Among 1092 donors, the allele with highest frequency were HLA-A*1101, A*0201, A*2402, A*0207, A*3303, A*0206 and A*3001; HLA-B*4001, B*4601, B*5801, B*1302, B*1501, B*5101and B*1301; HLA-Cw*0102, Cw*0702, Cw*0304, Cw*0801, Cw*0602, Cw*0303, Cw*0302 and Cw*0401; HLA-DRB1*0901, DRB1*1501, DRB1*1202, DRB1*0701, DRB1*0803, DRB1*0405, DRB1*0301 and DRB1*1101; HLA-DQB1*0301, DQB1*0303, DQB1*0601, DQB1*0202, DQB1*0602, DQB1*0302, DQB1*0401, DQB1*0201 and DQB1*0502. The probability of HLA high-resolution DNA matching between 1092 donors and 931 patients(10/10 match) was 16.7%. Mismatching at a single HLA-A, -B, -C, -DRB1 or DQB1 locus (9/10) was 17.7%. A single mismatch at each locus of HLA-A, -Cw,- DRB1,- DQB1,- B was 6.8%, 6.3%, 2.0%, 1.7%, and 0.8%, respectively. Double mismatch (8/10) was 18.4%, such as loci A+ Cw(5.0%), DRB1+DQB1(4.6%) and B+ Cw(3.8%). The donor/patient pairs mismatched between allele of A*0201 and A*0206, A*0201 and A*0207, A*1101 and A*1102, B*4006 and B*4002, B*1501 and B*1527, Cw*0304 and Cw*0302, Cw*0304 and Cw*0303, DRB1*1501 and DRB1*1502, DRB1*1202 and DRB1*1201, DRB1*0406 and DRB1*0403, DRB1*1401 and DRB1*1454, DQB1*0303 and DQB1*0302, respectively, were statistically associated with lower-risk Allo-HSCT. These results suggested that high-resolution DNA matching or mismatching for HLA-A, -B, -C, -DRB1 and DQB1 alleles could be associated with better clinical outcome and higher survival. Furthermore, the identification of high risk mismatch and permissive mismatch would be beneficial for the selection of a suitable donor.

Disclosures:

No relevant conflicts of interest to declare.
Severe Chronic Neutropenia in Korean Children.
Authors: Lee Mee Jeong , Park Hyeon Jin , Shin Hee Young , Kang Hyoung Jin , Koo Hong Hoe , Kook Hoon , Kwon Young Joo , Kim Sun Young , Kim Soon Ki , Kim Heung Sik , Moon Hyung Nam , Park Kyung Duk , Park Byung-Kiu , Park Seong Shik , Park Eun Sil , Baek Hee Jo , Seo Jong Jin , Sung Ki Woong , Song Joon Sup , Ahn Hyo Seop , Won Sung Chul , Yoo Keon Hee , Lyu Chuhl Joo , Lee Kun Soo , Lee Soo Hyun , Lee Young Ho , Lim Yeon Jung , Lim Young Tak , Lim Jae-Young , Im Ho Joon , Han Dong kyun , Hwang Tai Ju
Published: October 2019

Abstract 4513

Introduction

Severe chronic neutropenia (SCN) is a rare hematologic disorder defined by an absolute neutrophil count less than 0.5×109/L for several months or years. They usually suffer from recurrent infections. Principal subtypes of SCN are congenital, cyclic, idiopathic and primary autoimmune neutropenia (AIN).

Patients and Methods

Medical records collected from a national survey were retrospectively analyzed on newly diagnosed SCN patients in Korea between January, 1999 and December, 2008 in respect to the diagnosis, clinical manifestations, treatments and prognosis of the patients.

Results

There were 64 patients (Male, 20; Female 44) reported from 16 hospitals: congenital, 19; cyclic, 16; idiopathic, 25; and immune in origin, 4. The main clinical manifestation was various types of bacterial infections. Two cases (1 congenital, 1 cyclic) were diagnosed by family histories. The median age at diagnosis was 12 months (11 days-158 months). A bone marrow examination was done in 45 patients (70.3 %) at the median age of 26 months (1 day-158 months), with the interval between the initial CBC and BM study being 7.3 months (9 days-138 months). The ELA2 mutation, done in 6 patients, was not detected. Only one patient with congenital SCN evolved to AML at 54 months after diagnosis, who is under chemotherapy. Most patients were treated with G-CSF (5-10 μg/kg/day) during infection episodes. The median follow up duration was 23 months (11 days – 176 months). Two patients of congenital SCN died of infection (pneumonia, meningitis) and 8 patients were lost to follow up, and the remaining are alive.

Conclusions

SCN is a rare hematologic disease with inherent vulnerability to infections, thus early detection with proper management should be important for survival of SCN patients. We propose a nation-wide, prospective study to delineate the prevalence, molecular diagnosis, natural history, the optimal use of G-CSF, and prognostic factors in Korean patients with SCN.

Disclosures:

No relevant conflicts of interest to declare.
Expected Long-Term Survival of Older Patients Diagnosed with Non-Hodgkin Lymphoma in 2007-2011.
Authors: Pulte Dianne , Gondos Adam , Brenner Hermann
Published: October 2019

Abstract 4525

Background

New therapeutic options have led to substantial increases in survival expectations of patients with non-Hodgkin lymphoma (NHL) in recent years. In contrast to many malignancies, survival in older patients has improved in NHL at a rate similar to that in younger patients. In the past, the impact of these innovations on long-term survival of NHL patients on the population level has only been disclosed with substantial delay. In order to reduce this delay, we employed a newly developed projection method to estimate survival of patients age 60 and older with NHL in 2007-11.

Methods

In order to demonstrate the validity of model based projection, we calculated survival for 4 prior periods for which observed 5-year survival data is available by three methods: cohort analysis, period analysis, and model based projection and compared the results to the actual observed survivals from the same time periods. We next calculated survival estimates for the most recent patient cohort for which 5- and 10-year survival data is available, period analysis for 2002-06, and projection estimates for 2007-11.

Results

A preliminary empirical evaluation of the method using historical data indicated good performance in projection of age specific and overall 5- and 10-year relative survival in older patients with model based projection giving a result closer to the observed survival than either cohort analysis or period analysis for all age groups in each time period. Five and 10-year survival estimates for 2007-11 for patients aged 60+ were 65.1% and 53.5%, respectively, 8.9 percentage units (% units) and 14.8% units, respectively, higher than survival estimated from the most recent cohort analysis available (see table). Age specific 5- and 10-year relative survival estimates using model based projection ranged from 76.9% and 66.6%, respectively, for age 60-64 to 50.7% and 37.5%, respectively, for patients age 80+. Survival estimates by model based projection were higher for all age groups including 80+, both high and low grade disease, nodal and extranodal disease, and both genders.

Conclusions

Patients over 60 diagnosed with NHL in 2007-2011 have much higher long-term survival expectations than suggested by previously available survival statistics. The use of model based projection is reliable in this patient population and demonstrates steady improvement in survival over time for older patients with NHL. This reinforces the findings from clinical trials which show that treatment of NHL is tolerable and effective in older patients.

Disclosures:

No relevant conflicts of interest to declare.