bluuebunny/crossref_metadata_2025 huggingface dataset:
Outcome Comparison of Lymphoma and Myeloma Patients after Autologous Stem Cell Transplantation (ASCT) with Peripheral Blood Stem Cell Mobilization Between Plerixafor (P) Mobilized in Poor Mobilizer Patients and Non-Plerixafor Mobilized Patients
Authors: Malik Shahbaz A. , Yanovich Saul , Rapoport Aaron P. , Badros Ashraf Z , Hardy Nancy M. , Kocoglu Mehmet H. , Duffy Alison , Ruehle Kathleen , Nishioka Jennifer , Goloubeva Olga , Yared Jean
Published: October 2019

Background: P is used in combination with G-CSF to improve the mobilization of peripheral blood hematopoietic stem cells in poor mobilizers. Limited data are available in regard to effects of P on post-transplant outcomes in comparison with non-P chemomobilized patients.

Methods: In this retrospective study, we compare the engraftment and the outcomes of 34 patients mobilized with P + G-CSF or just-in-time rescue P in combination with chemotherapy and G-CSF to 143 patients (control group) mobilized with G-CSF with or without chemotherapy in lymphoma and myeloma patients who underwent ASCT between February 2012 and April 2014 at the University of Maryland Greenebaum Cancer Center. Post-transplantation outcomes including infections, hematologic recovery, relapse, progression and survival were recorded.

Results: The median number of collected of CD34+ cells/Kg was 5.9 x 10(6) in the P group and 12.3 x 10(6) in the control group (p=0.0002). Median time to neutrophil engraftment (>0.5 × 10(9) /L) was comparable between the 2 groups: 12 days for the P group and 11 for the control group (p=0.28). There was a trend toward a shorter time to platelet engraftment (>20 × 10(9) /L) in the control (12 days) compared to the P group (14 days) (p=0.056). Progression-free survival at 1 year after (ASCT) was 88.2% in the P group and 81.8% in the control group. There was no difference in the overall survival of both groups (p=0.62)

Conclusions: Short and long-term engraftment and outcomes after ASCT seem to be comparable in lymphoma and myeloma patients receiving plerixafor compared to chemomobilized patients without plerixafor. This observation support the use of plerixafor + G-CSF or just-in-time rescue P in patients who mobilize poorly without P.

Figure 1. Figure 1.

Disclosures

No relevant conflicts of interest to declare.
Effect of Body Mass Index on Overall Survival of Patients with Allogeneic Hematopoietic Stem Cell Transplantation
Authors: Xue Shengli , Yang Jing , Zhang Xiang , Shen Yueping , Depei Wu
Published: October 2019

AIM: The present study was performed to investigate the association between body mass index (BMI) before transplantation and the overall survival (OS) of patients with allogeneic hematopoietic stem cell transplantation (allo-HSCT).

METHODS: Data from 310 adults who were diagnosed with acute leukemia and underwent allo-HSCT between March 2001 and December 2011 were analyzed. Demographic and clinical data were collected from medical records. In accordance with the suggested BMI categories for Asian population, patients with BMIs of ≥23 and ≥25 kg/m2 were identified as overweight and obese, respectively. Hazard ratios (HRs) with 95% confidence intervals (CIs) were estimated using Cox proportional hazards models, which were adjusted for possible confounding factors.

RESULTS: The median follow-up time among the patients was 19.7 months (interquartile range = 8.1 to 37.7). A total of 93 (34.8%) patients died within the follow-up period. After adjusted for the potential confounders, normal-weight, overweight, and obese patients showed significantly lower HRs than underweight patients, with a significant trend of OS improvement upon increasing BMI (P = 0.019). Overweight and obese patients survived longer, with a significantly decreased HR by approximately 40% (HR = 0.60;95% CI: 0.38 to 0.95) compared with underweight and normal-weight patients.

CONCLUSIONS: An increased OS was seen in allo-HSCT patients with BMI≥23 compared to those with lower BMI. Further studies are required to obtain better understanding of the effects of BMI or body composition on the survival of allo-HSCT patients.

Disclosures

No relevant conflicts of interest to declare.
A Novel Compound Inhibits Chronic Myeloid Leukemia Via Upregulating Apoptosis
Authors: Xin Jiajia , Yin Dandan , Fu Wei , Zhang Hui-Jie , Chen Yaozhen , Wang Yazhou , Li Mingkai , Hu Xingbin
Published: October 2019

Chronic myeloid leukemia (CML) is a myeloid proliferative disorder mainly result from chimeric protein BCR-ABL1 encoded by a fusion gene at the t(9;22) (q34;q11) chromosomal translocation. Intrinsically, this recombined protein results in an increased tyrosine kinase (TK) activity that directly related to hematopoietic stem cell malignant proliferation. Consequently, the drugs derived from tyrosine kinase inhibitors (TKI) have been developed as an infective therapy, and greatly improved patients survival in clinic. Unfortunately, single TKI administration led to toxicities or tolerance in long-term treated CML patients. Even worse is, about 5% CML patients were not caused by bcr-abl gene mutation. Thus better medicines are badly needed to compensate CML therapy. Herein, we investigated the undefined function of a biscoumarins. The new synthesized compound exhibited a null toxicity on HUVECs but intensive toxicity on K562 leukemic cells. Subsequent results demonstrated that it efficiently inhibited the expansion of human CML cell line and bone marrow cells of SCL-tTA-BCL/ABL transgenic model mice via increased apoptosis. Critically, we also showed that CD34+ bone marrow leukemic cells collected from patients underwent more apoptosis after treated by the biscoumarins derivate. To extend these results into vivo, we observed a prolonged survival of bcr-abl transgenic mice treated by derivate mono-therapy or combination with imatinib compared to those of untreated or imatinib-treated CML mice. All together, these results indicated that this biscoumarins derivate may have novel potential as a therapeutic agent against CML.

Disclosures

No relevant conflicts of interest to declare.
Blastic Plasmacytoid Dendritic Cell Neoplasm: A Single-Center Experience Documenting Frequent CNS Involvement
Authors: Eulitt Patrick , McNally Gretchen A. , Porcu Pierluigi , Klisovic Rebecca B. , Vasu Sumithra , Baiocchi Robert A
Published: October 2019

Blastic Plasmacytoid Dendritic Cell Neoplasm (BPDCN) is a rare myeloid malignancy that often manifests with skin, lymph node, and bone marrow involvement. Patients diagnosed with BPDCN have a poor prognosis, with a median overall survival ranging from 12-14 months (Julia et al., 2014). Aoki et. al (2015) published a retrospective review reporting overall survival at 4 years for patients who underwent autologous and allogeneic hematopoietic stem cell transplant (HSCT) as 82% and 53%, respectively. Progressive disease, including central nervous system (CNS) involvement, excludes patients from receiving HSCT. Published case reports suggest that CNS presentation at diagnosis is rare, with estimates of only 10% of patients with CNS involvement at diagnosis (Feng et al., 2014; Starck et al., 2014). A retrospective case series of all patients treated for BPDCN at The Ohio State University from 1990 to present (n=11) was performed. Demographic data is summarized in table 1. Four of five patients undergoing CNS evaluation at diagnosis were found to have CNS involvement. All eleven patients underwent chemotherapy with the majority receiving induction chemotherapy (table 1). Three of the four patients with known CNS involvement at diagnosis expired within three months. The fourth patient survived with significant neurological complications secondary to intrathecal chemotherapy and was not a HSCT candidate. Two of eleven patients underwent HSCT. One died from relapsed disease in bone marrow, and one achieved long-term progression free survival. Two patients were lost to follow up, and were 7 and 16 months past diagnosis at last contact. CNS involvement with BPDCM at diagnosis may be more common than the literature suggests and should be investigated at diagnosis to allow these patients the best chance of long-term survival.

Table 1. Patient characteristics and treatment summary Mean age at diagnosis 61.4 years Stage at diagnosis IV: 81.8% I: 18.2% BM involvement at diagnosis Yes: 81.8% No: 18.2% CNS involvement at diagnosis Yes: 36.4% No: 9.1% Unknown: 54.5% CNS involvement during clinical course Yes: 63.7% No: 18.2% Unknown: 18.2% Chemotherapy Hyper-CVAD: 45.5% CHOP: 18.2% EPOCH: 18.2% Clinical Trial: 18.2% SCT Yes: 18.2% No: 72.7% Unknown:9.1% Survival < 12 months: 45.5% 12-24 months: 27.3% >24 months: 18.2% Lost to follow up: 18.2% Median time to death in months 10.0* (range 1-26) *Time to death calculation excluded two patients lost to follow-up and two patients that are still living

Disclosures

Porcu: Cell Medica: Research Funding; Infinity: Research Funding; Seattle Genetics: Research Funding; Shape: Research Funding; Celgene: Research Funding.
EGFR Signaling in Osteoblasts Regulates Circadian Rhythm of HSPC in Circulation
Authors: Kumar Sachin , Vassallo Jeff , Nattamai Kalpana , Cancelas Jose A , Geiger Hartmut
Published: October 2019

EGFR signaling regulates growth, differentiation, proliferation and migration in multiple organ systems. We previously demonstrated that inhibition of EGFR signaling on hematopoietic stem and progenitor cells (HSPCs) enhances G-CSF induced stem cell mobilization, while preliminary data suggested that inhibition of EGFR signaling in the stem cell niche actually had the opposite effect of inhibiting mobilization (Ryan et al., Nat Med, 2010). We thus tested the novel hypothesis that there is a role for EGFR signaling in the bone marrow (BM) niche with respect to regulating hematopoiesis. We utilized a set of mouse strains that express the recombinase Cre under distinct promoters to specifically delete EGFR in various types of stroma cells including Col-Cre; EGFRf/f (deletion in osteoprogenitor/osteoblasts (OBs)), Dermo-Cre; EGFRf/f (mesenchymal stem cells (MSCs) including chondrocytes and OB), Tie2-Cre; EGFRf/f (Endothelial cells) and Nestin-Cre; EGFRf/f (Schwann/neural cells) and compared them to no Cre-EGFRf/f mice as control wild type for EGFR.

Basic parameters of steady-state hematopoiesis were not altered in mice devoid of EGFR signaling in the various types of stroma cells listed above. We further investigated HSPC mobilization in EGFRf/f mice and interestingly, Col-cre and Dermo-cre EGFRf/f mice exhibited a lower number of circulating HSPC in blood in comparison to wild type mice, as determined by colony forming units (CFUs) or flow cytometry. Deletion of the EGFR in endothelial (Tie2-Cre) and neuronal (Nestin-Cre) compartments did not result in a decline in the number of circulating HSPCs. Upon G-CSF challenge, Col-cre and Dermo-cre EGFRf/f mice mobilized HSPCs similar to controls, suggesting that EGFR signaling in OBs/MSCs is dispensable for G-CSF induced mobilization. HSPCs circulation under steady state follows a circadian rhythm. We next tested whether EGFR signaling in OBs might play a role in circadian rhythm driven HSPC circulation. After 5 hours of light cycle i.e. Zeitgeber time-5 (ZT-5), when the number of circulating HSPCs is high, Col-Cre EGFRf/f and Dermo-Cre EGFRf/f mice presented with low numbers of circulating HSPCs. After 13 hours of light cycle (ZT-13) (low number of circulating HSPCs), the number of HSPCs in blood in Col-Cre EGFRf/f and Dermo-Cre EGFRf/f mice were similar to controls. Together, this suggests that EGFR signaling in OBs is essential for the rhythmic increase in circulating HSPC in blood at ZT5. Currently, we are investigating molecular mechanisms driven by EGFR signaling in OBs that regulate HSPC retention in BM and circadian regulation of EGFR signaling. In summary, our data suggest an important and also very specific (no other phenotype altered) role of EGFR signaling for regulating the circadian rhythm of HSPC circulation in peripheral blood.

Disclosures

No relevant conflicts of interest to declare.
New CNV Regions Identified in ITP Provide Evidence for Genetic Predisposition
Authors: Ar M. Cem , Yucesan Emrah , Yalniz Firat , Hatirnaz Ng Ozden , Salihoglu Ayse , Berk Selin , Eskazan Ahmet Emre , Ongoren Seniz , Baslar Zafer , Ozbek Ugur , Soysal Teoman , Aydin Yildiz , Sayitoglu Muge
Published: October 2019

Immune thrombocytopenia (ITP) is an autoimmune disease resulting from inhibition of megakaryopoiesis and destruction of platelets by platelet reactive autoantibodies. Adult-onset ITP usually exhibits a relapsing chronic course and is often associated with other disorders of autoimmune or infectious origin, including systemic lupus erythematosus (SLE), lymphoproliferative diseases, common variable immunodeficiency (CVID) disease, and human immunodeficiency virus (HIV) infection. The exact pathophysiology of ITP has not been fully elucidated. Heterogeneity in disease presentation and in response to immunosuppressive therapy suggests different underlying mechanisms; and thus justifies the investigation of relevant genetic associations. Here we tried to unravel the genetic background of the immune dysfunction in ITP by using SNP array technology.

We analyzed the blood samples of 12 adults (6 males and 6 females) with primary refractory ITP, by SNP array approach to identify candidate genomic regions. None of the patients showed clinical and laboratory findings of autoimmune diseases, malignancy or infections. All individuals were genotyped using Illumina Human HumanCytoSNP-12 BeadChip (300K). Whole genome SNP genotyping data were delineated using GenomeStudio software platform. The genotyping data was obtained by further analyzes in terms of copy number variation (CNV).

In total we observed 14 CNVs in 12 patients. All CNVs were on autosomal chromosomes; 8 duplications and 6 deletions (gain 2p, 9q, 13q, 14q, 17p, 17q, 20q; loss 5q, 7p, 7q, 15q, 17q, 19q). The rearrangements sizes were between 45.42 Mb and 1.08 Mb. Moreover in five patients the duplications were in the form of mosaic structural genomic rearrangements. These mosaic duplications were found at 17q21.31 (0.2 Mb), 2p23.2 (0.34 Mb), 9q21.2 (10.89 Mb), 9q31.2 (17.77 Mb), 14q13.2 (0.37 Mb), 17p12 (1.46 Mb), 20q11.21 (14.24 Mb). None of the detected CNV regions could be demonstrated in two independent cohorts of 30 individuals with non-hematological disorders and healthy controls.

Little is known on the genetic background of immune thrombocytopenia. Our aim was to investigate the impact of CNVs on the pathophysiology of ITP. We found several gains/losses in different chromosomal regions. However, the most striking result of our study was the detection of mosaic patterns that have been reported to most commonly associate with hematological cancers. These mosaic patterns may be associated with clonal expansion of T- or B-cells, leading to the immune dysfunction seen in ITP. Evaluation of the detected regions and their relation to B and T- cell clonality is ongoing. These preliminary results provide early evidence for the presence of predisposing CNVs in adult onset ITP.

Disclosures

No relevant conflicts of interest to declare.
Interleukin-1 Drives Precocious Myeloid Differentiation of Hematopoietic Stem Cells at the Expense of Self-Renewal
Authors: Pietras Eric Martin , Mirantes-Barbeito Cristina , Fong Sarah , Loffler Dirk , Kovtonyuk Larisa Vladimirovna , Lakshminarashimhan Ranjani , Chin Chih Peng , Techner Jose-Marc , Will Britta , Nerlov Claus , Steidl Ulrich , Manz Markus , Schroeder Timm , Passegue Emmanuelle
Published: October 2019

Hematopoietic stem cells (HSCs) maintain lifelong blood homeostasis. While many of the cell-intrinsic mechanisms regulating HSC function at steady state have been well characterized, the role of inflammatory cytokines and other environmental factors in tailoring blood production following physiological insults has become a topic of emerging interest. The cytokine interleukin-1 (IL-1) is a prototypical pro-inflammatory cytokine that plays a key role in host inflammatory responses to injury and infection, and is associated with elevated myeloid cell production. Importantly, IL-1 also drives a wide range of chronic inflammatory conditions such as diabetes, obesity, and arthritis that are often characterized by deregulated blood homeostasis. Here, we show at single-cell resolution using continuous tracking technology that IL-1 drives accelerated HSC cell division kinetics and myeloid differentiation via the rapid activation of a precocious PU.1-dependent myeloid gene program. Activation of this program requires direct IL-1R signaling and subsequent activation of IKK kinases, and instructively primes HSCs to adopt a myeloid fate. Strikingly, we demonstrate that IL-1 produced by myeloid cells and endothelial cells of the bone marrow (BM) niche exerts similar effects in vivo, and is required for efficient myeloid recovery following acute challenges such as transplantation or myeloablation. On the other hand, we find that chronic IL-1 exposure substantially remodels HSC blood output, resulting in myeloid overproduction and expansion of myeloid-biased multipotent progenitor (MPP) compartments at the expense of lymphoid and erythroid lineages. Critically, chronic IL-1 erodes HSC self-renewal, significantly impairing their regenerative capacity following transplantation. On the other hand, chronically exposed HSCs recover their function upon IL-1 withdrawal. Collectively, these findings identify IL-1 as a critical regulator of HSC fate and lineage specification via activation of a PU.1 circuit. They also demonstrate a role for IL-1 as a double-edged sword in HSC biology, promoting HSC regeneration in response to acute insults while severely disrupting HSC self-renewal and lineage output during chronic exposure, hence identifying IL-1 as an important and therapeutically targetable factor underwriting myeloid overproduction and other deregulations that contribute to the pathogenesis of a variety of chronic inflammatory diseases and blood disorders.

Disclosures

No relevant conflicts of interest to declare.
Tumor Suppressors BTG1 and IKZF1 Cooperate during Mouse Leukemia Development and Impact Relapse Rate in Childhood Acute Lymphoblastic Leukemia
Authors: Scheijen Blanca , Tijchon Esther , van Ingen Schenau Dorette , Marke Rene , van Emst Liesbeth , Van Der Meer Laurens T , Kuiper Roland P , Boer Judith M. , Pieters Rob , Hoogerbrugge Peter M , Den Boer Monique L. , van Leeuwen Frank N
Published: October 2019

Deletions and mutations affecting transcription factor IKZF1 are associated with increased relapse risk and poor outcome in B cell precursor acute lymphoblastic leukemia (BCP-ALL). However, additional genetic events may either enhance or negate the effects of IKZF1 on prognosis. We observed that deletions of the gene encoding the transcriptional coregulator BTG1 frequently co-occured with loss of IKZF1 function, suggesting a synergistic role for these events during leukemia development or progression. Targeted deletion of Btg1 predisposed both Btg1+/- and Btg1-/- mice to T cell malignancies, similar to what has been observed in Ikzf1 heterozygous knockout animals. Hence, while somatic single single-copy losses of either BTG1 and IKZF1 in the patient are predominantly found in BCP-ALL, targeted deletion of these genes in the mouse gives rise to T cell malignancies.

To establish whether loss of BTG1 function affected the tumor suppressive role of IKZF1, the Btg1 knockout allele was crossed onto mice heterozygous for a loss-of-function Ikzf1 allele. Leukemia penetrance in these compound mice increased in a Btg1 dose-dependent manner. These leukemias were characterized by clonal TCRb rearrangement and aggressive infiltration into secondary organs, indicating synergistic roles for these tumor suppressors during mouse leukemia development.

To investigate the effects of combined IKZF1/BTG1 loss in human BCP-ALL, we examined a large pediatric cohort of BCP-ALL cases, and found that the combined presence of BTG1 and IKZF1 deletions was associated with a markedly higher incidence of relapse, relative to IKZF1-deleted cases without BTG1 aberrations. Similar to BTG1 copy number losses, deletions in EBF1, PAX5, RB1 and CDKN2A/B appeared to be selectively enriched in IKZF1 deleted ALL. However, in contrast to BTG1, none of these other copy number alterations affected relapse incidence or outcome in this patient group. In conclusion, our data demonstrate synergy between the tumor suppressors BTG1 and IKZF1 during mouse leukemia development while the combined (single copy) loss of these two tumor suppressors identifies a patient group with an extremely poor outcome.

Event free survival in a cohort of 514 children newly diagnosed with BCP-ALL, divided into four categories based on IKZF1 and BTG1 deletion status.

Figure 1. Combined loss of IKZF1 and BTG1 predicts poor outcome in BCP-ALL Figure 1. Combined loss of IKZF1 and BTG1 predicts poor outcome in BCP-ALL

Disclosures

Pieters: Eusa Pharma: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees.
Hepcidin and Atoh8 mRNA Expression Are Decreased in Murine Models of Hemochromatosis However Hepcidin Expression Is Unaffected By Atoh8 Knockout
Authors: Fleming Robert E , Li Yihang , Miller Ian , Prasad Princy , McKie Andrew
Published: October 2019

Atoh8 is a member of the bHLH transcription factor family, and hepatic expression of atoh8 mRNA is up-regulated by dietary iron and down-regulated by stimulation of erythropoiesis in mice, in parallel with changes in hepcidin mRNA(hamp1). It is a candidate transcription factor in the regulation of Hamp1 by iron status. Here we performed studies evaluating the contribution of genes regulating hepcidin to the regulation of Atoh8, as well as the role of Atoh8 in the regulation of hepcidin. In the former studies, wild type FVB and three hereditary hemochromatosis (HH) mouse models, Hfe-/-, TfR2Y245X/Y245X and Hfe-/- TfR2Y245X/Y245X mice, were placed on diets containing 60 ppm or 25,000 ppm iron at weaning for two weeks before euthanasia and tissue collection. Liver expression of Hamp1 and Atoh8 were evaluated. Additionally, the WT and HH mice were intraperitoneally injected with 30 mg/kg phenylhydrazine (PHZ) on days 49 and 50, and then euthanized on day 53. Liver and spleen non-heme iron concentrations, and serum hematologic and iron parameters were measured. The liver mRNAs for hamp1, atoh8, and bmp6 were measured by quantitative RT-PCR. Hfe-/- and Tfr2Y245X/Y245X mice each demonstrated lower Hamp1 and Atoh8 expression than did WT mice, despite hepatic iron loading. WT mice demonstrated the expected increases in Hamp1 and Atoh8 expression in response to dietary iron loading. Hamp1 increased in the HH mice with iron loading; however Atoh8 expression was not statistically different. The addition of Hfe knockout to the TfR2Y245X/Y245X mutation suppressed Hamp1 expression further, but had no additive effect on Atoh8. PHZ treatment at 30 mg/kg mouse significantly decreased red blood cells and hematocrit, caused severe splenomegaly, and down-regulated Hamp1 and Atoh8 in WT mice, as well as in HH mice. These results are consistent with a role for Hfe and TfR2 in normal signaling to Hamp1 and Atoh8, but also suggest some regulation of Hamp1 in response to dietary iron independent of a change in Atoh8 expression. To evaluate the role of Atoh8 in the regulation of hepcidin, Atoh8 Δexon1 homozygous, heterozygous and WT mice were placed on diets containing standard (250 ppm) and euthanized at 5-6 weeks of age. The Atoh8 Δexon1 homozygous mice were significantly smaller in size than the WT or heterozygous littermates, but did not differ in any measured iron parameter (serum iron, hematocrit, liver iron, liver hamp1 mRNA, liver bmp6 mRNA). The Atoh8 Δexon1 homozygous mice also did not differ from heterozygous mice in PHZ response. These observations indicate that HFE and TFR2 are necessary for normal Atoh8 as well as hepcidin expression; however they do not support an essential role for Atoh8 in hepcidin regulation by iron demand.

Disclosures

No relevant conflicts of interest to declare.
Resolvin D1 and Resolvin D2 Protect Against Hypoxia/Reoxygenation Induced Lung and Kidney Damage in a Sickle Cell Mouse Model of Acute Vaso-Occlusive Crisis
Authors: Mattè Alessandro , Recchiuti Antonio , Serhan Charles , Siciliano Angela , Federti Enrica , Weinberg Olga , Kalish Brian , Brugnara Carlo , De Franceschi Lucia
Published: October 2019

Sickle cell disease (SCD) is characterized by hemolytic anemia in association with acute and chronic life-threatening clinical complications. Acute vaso-occlusive crisis (VOCs) are the main cause of hospitalization for SCD patients. In VOCs, amplified inflammatory response plays a key role in acute organ damage. Pro-Resolving lipid mediators such as resolvins (Rv) accelerate resolution of acute inflammation in different models, indicating that stimulation of endogenous resolution of inflammatory processes may be an additional strategy in limiting tissue damage. Recent data suggest beneficial effects of Rvs in hypoxia/reoxygenation (H/R) related tissue injury. Here, we study the effects of Rvs on a model of acute VOCs using humanized SCD mice (Hbatm1(HBA)Tow Hbbtm2(HBG1,HBB*)Tow). We treated SCD and control healthy mice (AA, Hbatm1(HBA)Tow Hbbtm3(HBG1,HBB)Tow) (n =6-7 animals in each group) with RvD1, 7S, 8R, 17S- trihydroxy-docosa-4Z, 9E, 11E, 13Z, 15E, 19Z-hexaenoic acid, 17R- RvD1, 7S, 8R, 17R- trihydroxy-docosa-4Z, 9E, 11E, 13Z, 15E, 19Z-hexaenoic acid; RvD2, 7S, 16R, 17S-trihydroxy-docosa-4Z, 8E, 10Z, 12E, 14E, 19Z-hexaenoic acid. Mice were treated with RvD1 and RvD2 at the dose of 2.5 ug/Kg by gavage 1 hour (hr) before H/R stress (10 hrs 8% oxygen followed by 3 hrs reoxygenation), which we have used in the past to mimic acute VOCs. We found that RvD1 and RvD2 significantly reduced the H/R-induced (i) increase in neutrophil count; (ii) lung inflammatory cell infiltrate, mucus and thrombi formation; (iii) glomerular inflammatory cell infiltration, glomerular sclerosis and thrombi formation. In the lung of H/R SCD mice, RvD1 prevented the H/R induced up-regulation of (i) cytokines such as MCP2, IL-6 and ET-1; (ii) vascular endothelial activation markers (VCAM-1 and ICAM-1; (iii) cytoprotective systems such as Prx6 and HO-1. In the kidney of H/R SCD mice, RvD1 significantly reduced H/R induced expression of IL-6 and ET-1 as well as HO-1. Our data indicate that RvD1 and RvD2 modulating inflammatory responses related to H/R in SCD, protect sickle cell target organs, and foster resolution. Thus, RvD1 and RvD2 might represent a novel therapeutic approach for acute VOCs in SCD.

Disclosures

No relevant conflicts of interest to declare.
Placenta Growth Factor Links the IL-13 Response and the Leukotriene Pathway to Augment Airway Hyper-Responsiveness
Authors: Eiymo Mwa Mpollo Marthe-Sandrine , Brandt Eric , Shanmukhappa Shiva Kumar , Arumugam Paritha , Tiwari Swati , Loberg Anastacia , Rizvi Tilat , Lindsey Mark , Jonckx Bart , Carmeliet Peter , Kalra Vijay , Lecras Timothy , Ratner Nancy , wills-Karp Marsha , Khurana hershey Gurjit , Malik Punam
Published: October 2019

Airway hyper-responsiveness (AHR) affects 55-77% of children with sickle cell disease (SCD) and occurs even in the absence of asthma. While asthma increases SCD morbidity and mortality, the underlying mechanisms of this highly prevalent AHR in a hemoglobinopathy remain unknown. We hypothesized that Placenta Growth Factor (PlGF), an erythroblast-secreted factor that is elevated in SCD, mediates this AHR. We studied AHR in allergen-exposed PlGF-/- and SCD mice. Sickle mice genetically deficient in PlGF were not viable. PlGF augmented AHR, lung inflammation and blood/lung eosinophilia via IL-13 (a Th2-cytokine) and increased 5-lipoxygenase (a leukotriene synthetic enzyme) expression. AHR, or pulmonary inflammation and leukotriene levels were blunted in PlGF-/- mice or in PlGFWT mice treated with anti-PlGF-Ab, and was rescued by intratracheal administration of leukotrienes to PlGF-/- mice. Notably, Th2-mediated Stat-6 activation further increased PlGF expression from lung epithelium, eosinophils and macrophages, linking the PlGF-leukotriene pathway in a positive feedback loop. Th2 response is classically seen in asthma, and indeed, expression of PlGF and its downstream genes was increased in respiratory epithelial cells of asthma patients. Like patients with SCD, sickle hematopoietic chimeras developed increased AHR and had higher leukotriene levels; and both were abrogated by anti-PlGF-Ab or zileuton (5-lipoxygenase inhibitor) phenocopying the blunted AHR in PlGF-/- mice, and indicating that sickle erythroid-cells/RBC potentiate AHR. Overall, PlGF exacerbates AHR and uniquely links the leukotriene and Th2 pathways in asthma. Both zileuton, licensed for asthma and anti-PlGF-Ab, in clinical trials for cancer, could be promising therapies to reduce the pulmonary morbidity in SCD.

Disclosures

Jonckx: ThromboGenics: Employment.
Factor XIII and Red Blood Cells in Venous Thrombosis
Authors: Wolberg Alisa S.
Published: October 2019

Blood coagulation culminates in the thrombin-mediated conversion of fibrinogen to fibrin and production of a fibrin network that stabilizes the clot. The transglutaminase factor XIII(a) [FXIII(a)] crosslinks fibrin and increases the clot's resistance to fibrinolysis and mechanical disruption. Consequently, fibrin formation and crosslinking are essential for hemostasis. However, the formation of clots with abnormal fibrin network structure and/or stability (resistance to mechanical disruption or pharmacological dissolution) is a risk factor for both arterial and venous thrombosis. Improved understanding of fibrin formation and crosslinking during coagulation is essential for understanding the pathophysiologic mechanisms that promote thrombotic disease. We have shown that elevated levels of fibrinogen accelerate vessel occlusion and enhance thrombus stability in mice, demonstrating etiologic contributions of fibrin(ogen) to thrombosis. Recently, we discovered that FXIIIa transglutaminase activity also mediates venous thrombosis via its ability to promote red blood cell retention during clot contraction in whole blood. Genetic deletion of FXIII or pharmacologic inhibition of FXIIIa activity reduces red blood cell retention in clots and consequently, reduces thrombus size in vitro and in vivo. Furthermore, FXIII zymogen binding to soluble, circulating fibrinogen is necessary for normal FXIII activation and activity. The fibrinogen motif that mediates FXIII binding involves C-terminal residues in the fibrinogen gamma chain (gamma 390-396); mice that express a variant fibrinogen that has mutations within these residues show reduced binding of FXIII zymogen to fibrinogen, and delayed activation of FXIII. In venous thrombosis models, these mice phenocopy FXIII-deficient mice, producing smaller venous thrombi with reduced red blood cell content compared to thrombi from wild-type mice. Intriguingly, these mice do not exhibit bleeding or poor wound healing normally associated with FXIII deficiency. Collectively, these findings provide new insight into the pathophysiology of venous thrombosis, and expose the fibrin(ogen)-FXIII axis as a central determinant of venous thrombus formation and composition.

Disclosures

No relevant conflicts of interest to declare.
Choice of Better T-cells for Adoptive Immunotherapy
Authors: Busch Dirk Hans
Published: October 2019

Adoptive transfer of primary (unmodified) or genetically engineered antigen-specific T cells has demonstrated astonishing clinical results in the treatment of infections and some malignancies. The definition of optimal targets and antigen receptors as well as the differentiation status of transferred T cells are emerging as crucial parameters for generating cell products with predictable efficacy and safety profiles. Our laboratory has demonstrated that defined subsets within the memory CD8+ T cell compartment fulfill all key characteristics of adult tissue stem cells and are essential for robust and long-term maintained responses upon adoptive transfer. We have developed clinical multi-parameter enrichment technologies to purify these memory stem cells for clinical applications. In my presentation I will report on the status of ongoing clinical trials using such purified cell products either as a primary T cell population for the treatment of infections upon allogeneic stem cell transplantation or after genetic modification with a CD19 CAR for the treatment of malignancies (collaboration with Stan Riddell, FHCC/Seattle). Infusing small numbers of T cells within a memory stem cell product can be highly effective therapeutically, but bears some risk of toxicity. Therefore, safeguards that allow selective depletion of transferred cells in the case of un-tolerable side effects may be needed to further improve adoptive immunotherapy. I will present results exploring the capacity of a truncated version of EGFR (EGFRt) co-expressed with T cells expressing a CD19-CAR. In pre-clinical mouse models we demonstrate that application of Cetuximab, which binds to EGFRt, confers selective depletion of adoptively transferred CAR-T cells in vivo. Long-term B cell aplasia, which is a main side effect of CD19-CAR T cell therapy, can be completely reverted with this strategy. Vaccination studies upon B cell recovery demonstrate full functionality of antigen-specific antibody formation. EGFRt co-expressing CD19-CAR T cells have been successfully transferred into first human patients, providing the option to test for the first time in a clinical setting whether treatment of B cell aplasia after long-term leukemia remission can be achieved by selective depletion.

Disclosures

Busch: STAGE cell therapeutics: Other: I was share holder of STAGE cell therapeutics, a company that was recently bought by Juno therapeutics.. Off Label Use: CD19 CAR T cells.
Clonal Diversity and Mutated Genes in Myelodysplastic Syndromes
Authors: Ebert Benjamin L
Published: October 2019

Myelodysplastic syndromes (MDS) are driven by somatic genetic mutations that drive clonal explansion of neoplastic cells. The identification of these lesions has enabled large-scale studies to assess the association of genetic lesions with clinical and biological phenotype. Single mutations in driver genes in isolation, without concurrent mutations in additional driver genes, commonly do not cause overt disease, but rather cause an expanded population of blood cells without cytopenias, a condition that we have termed clonal hematopoiesis of indeterminate potential (CHIP). CHIP is common in the general population and is associated with age, risk of hematologic malignancy, and overall mortality. In contrast, patients with MDS generally have mutations in multiple genes as well as the disease-defining morphologic dysplastia and peripheral blood cytopenias. The particular mutations that drive MDS in an individual patient are powerfully associated with clinical phenotype, including cytopenias, specific morphologic abnormalities, blast percentage, overall survival, and response to therapy. Clonal architecture adds an additional layer of complexity as disease progression or response to therapy may be associated with shifts in the dominant genetic clone in the bone marrow. As the availability of genetic testing for panels of genes becomes widespread as a clinical assay, current endeavors are to move validated research findings into routine clinical use and to employ deep genetic analyses to evaluate response to investigational therapies in clinical trials.

Disclosures

Ebert: Genoptix: Consultancy, Patents & Royalties; H3 Biomedicine: Consultancy; Celgene: Consultancy.
The Role of the Tumor Microenvironment in Lymphoid Malignancies
Authors: Steidl Christian
Published: October 2019

Lymphoid cancers represent a heterogeneous group of neoplasms composed of malignant lymphoid cells with variable infiltration by non-neoplastic, mostly immune cells (tumor microenvironment). For some subtypes of lymphoid cancers, the contribution of the microenvironment to the histological appearance is widely recognized and used for pathological classification. Although microenvironment-related biology in lymphoid cancers has been primarily explored in a core set of lymphoma subtypes, the number of entities studied has recently accelerated. The pathogenic evolution of tumor microenvironments and in particular their composition and spatial distribution can be perceived as a complex function of 1) genetic alterations within the malignant cell population, 2) the extent and dependence on the molecular crosstalk involving cyto- and chemokines, and 3) host-specific factors. As a result, three major patterns of microenvironmental architecture can be distinguished termed "Re-education", "Recruitment" and "Effacement". Hodgkin lymphoma can serve as a paradigm for an extensive crosstalk between tumor cells and a quantitatively dominant tumor microenvironment. Importantly, related prognostic implications of tumor microenvironment composition (e.g. macrophage content, representation of T cell subsets) have been extensively studied in this disease. With focus on B cell lymphomas, this talk will highlight the emerging literature about genetic alterations in malignant lymphoma cells that provide the foundation for somatically acquired immune privilege and evasion from immune surveillance. The genomic changes discussed in this talk can be broadly categorized according to the effect that they exert on the tumor microenvironment: 1) Loss or down-regulation of (surface) molecules leading to decreased immunogenicity of tumor cells (e.g. mutations of B2M, CIITA); 2) Increased expression of surface molecules suppressing immune cell function (e.g. structural genomic changes of PDL1, PDL2); 3) Recruitment or induction of a regulatory cellular milieu (e.g. mutations in JAK-STAT and NFκB signaling pathways). The discovery of gene mutations underlying immune privilege, properties of the altered molecules, downstream functional consequences and clinical rationales for therapeutic intervention will be presented in the context of specific lymphoma subtypes. It will be discussed how precise description of genomic and molecular alterations underlying immune privilege might accelerate effective targeting of microenvironment-related biology in the clinical setting. Moreover, the development and clinical implementation of predictive biomarkers will be outlined that harbor the potential to inform on companion diagnostic approaches to accompany therapies such as immunological checkpoint inhibition.

Disclosures

No relevant conflicts of interest to declare.
Secondary Polycythemia and the Risk of Venous Thromboembolism (VTE) Among Hospitalized Patients in the United States
Authors: Giri Smith , Mehta Kathan Dilipbhai , Bhatt Vijaya R.
Published: October 2019

Introduction:The prevalence of secondary polycythemia is on the rise due to increasing prevalence of obesity, sleep apnea and other chronic cardiopulmonary diseases. While polycythemia vera is well known to increase the risk of VTE, the impact of secondary polycythemia on the development of VTE is less clear.

Methods: We utilized the Nationwide Inpatient Sample (NIS) database to identify all adults ≥18 years with secondary polycythemia (ICD-9-CM code 289.0) diagnosed during 2011. NIS is the largest all-payer inpatient database in the US that captures about 20% of all US hospitalizations. The outcome of interest was acute VTE identified using corresponding ICD codes for DVT or PE. We also identified the presence of the following confounding factors: malignancy, venous catheterization, long bone fracture, infection, trauma, mechanical intubation, congestive heart failure (CHF), pregnancy, length of hospital stay >5 days, stroke and prior history of VTE. Multivariate logistic regression was used to assess the impact of secondary polycythemia on the occurrence of VTE controlling the above confounding factors. Multicollinearity was assessed using variance inflation factor (VIF). All p-values were 2 sided, and the level of significance was chosen at 0.05. Statistical analysis was done using STATA 13.0 (StataCorp, College Station, TX) taking into account the complex sampling design of the database.

Results: A total of 3,972 hospitalizations with a primary or secondary diagnosis of secondary polycythemia were identified out of a total of 6,840,854 hospitalizations. Concurrent diagnosis of VTE was seen in 4.8% of patients with secondary polycythemia (n=190) as opposed to 2.3% of patients without secondary polycythemia (n=161,122) (odds ratio, OR 2.07; 95% confidence interval, CI 1.75-2.45; p value <0.01). In a multivariate analysis, secondary polycythemia continued to have a significantly greater risk of concurrent VTE (adjusted OR 1.87; 95% CI 1.58-2.22; p value <0.01) after adjusting for confounders including age, stroke, malignancy, infection, history of VTE, long bone fracture, trauma, CHF, pregnancy, mechanical intubation, nephrotic syndrome and hospital stay >5 days. No evidence of multicollinearity was noted in the final regression model (maximum VIF of 1.13).

Conclusion:

Within the limitations of the study design, our retrospective study indicates an independent association between secondary polycythemia and VTE in patients admitted in the hospital, however, this does not imply causality. Further, our study does not answer whether the use of therapeutic phlebotomy reduces the risk of VTE in patients with secondary polycythemia.

Disclosures

No relevant conflicts of interest to declare.
A Clinic-Genetic Score for Risk Assessment of Recurrent Venous Thrombo Embolism
Authors: Gerotziafas Grigorios T. , Onambele Manuella , Benzerara L , Mokrani H , Ketatni Hela , Planche V , Delassasseigne C , Elalamy Ismail
Published: October 2019

Background

Current risk assessment models to predict the risk of recurrent venous thromboembolism (VTE) in an individual patient (i.e Vienna Pedicitive Model, DASH score) include clinical and biochemical parameters and have showed discrimination values ranging from 0.64 to 0.68. Genetic variants associated to thrombophilia have showed to play a relevant role in predicting primary VTE. Combination of those genetic variables together with clinical parameters may improve the predictive capacity of current models for predicting recurrent VTE.

Aim

To analyze the predictive capacity of a new clinic-genetic score for predicting recurrence of VTE, Thrombo inCode-Recurrent (TiC-Reccurent).

Methods

A population of 55 patients with a first VTE and none or one or more recurrent VTE (21 males, 34 females; 47.8±14.7 years old) and 39 controls with only the first VTE event (13 males, 26 females; 40.2±13.2 years old) and the same follow-up was used.

A regression model was constructed including clinical variants (age, gender, diabetes status, pregnancy status, contraceptive hormonal therapy, smoker status and family history of VTE) and genetic variants (F5 rs6025/rs118203906/rs118203905, F2 rs1799963, F12 rs1801020, F13 rs5985, SERPINC1 rs121909548, and SERPINA10rs2232698 plus the A1 blood group (rs8176719, rs7853989, rs8176743, rs8176750).

The predictive capacity was assessed by calculating the c-statistic (AUC-ROC); sensitivity, specificity Positive and Negative Likehoood ratios and odds ratio of a positive test result.

Informed consent was obtained and the study was approved by the Institution Ethics Committee. This study complies with the guiding principles for experimental procedures found in the Declaration of Helsinkiof the World Medical Association.

Results

In an univariate analysis, none of the clinic or genetic variables showed a significant difference between the patient with or without recurrences. TiC-Recurrent score showed a predictive capacity for recurrent VTE measures as AUC-ROC: 0.738, p<0,0001. For TiC-Recurrent the clinical sensitivity was 81.82 and the specificity was 33.33. The predictive capacity measures as LR+, LR- were 1.23 and 0.55, respectively. The odds ratio for TiC-recurrent was 2.7.

Conclusions

TIC-Recurrent score showed a good predictive capacity to identify subject at high risk of suffering a recurrence of VTE. Although a direct comparison to current scores for predicting the risk for a recurrence of VTE was not possible in the present study if the observed predictive capability for TiC-recurrent is confirmed it will improve the capacity of current scores. Our study suggests that the use of a clinical-genetic algorithm could be an aid in the treatment of patient with a first VTE and contribute to the prevention of VTE recurrences.

Disclosures

No relevant conflicts of interest to declare.
Functional and Molecular Alterations of Bone Marrow Mesenchymal Stem and Progenitor Cells in Patients with Myelodysplastic Syndrome with Ring Sideroblast
Authors: Dolinska Monika , Xiao Pingnan , Johansson Anne-Sofie , Sandhow Lakshmi , Kondo Makoto , Bouderlique Thibault , Ungerstedt Johanna , Hellstrom-Lindberg Eva , Qian Hong
Published: October 2019

Myelodysplastic Syndrome with Ring Sideroblast (MDS-RS), a clonal hematopoietic cell neoplasm, is low risk MDS, characterized by anemia, hyperplastic ineffective erythropoiesis and marrow ring sideroblasts. Mouse studies have shown that bone marrow niche, including endothelial cells, osteoblasts, adipocytes and mesenchymal stem cells (MSCs), contribute to progression of various hematological disorders. However, in vivo contribution of the different bone marrow stromal cells to the progression of MDS-RS in patients remains largely unknown.

To investigate this, we have phenotypically, molecularly and functionally characterized the BM native stromal cell subsets including MSCs freshly isolated by multi-color fluorescence activated cell sorting (FACS) from bone marrow of MDS-RS patients and age-matched healthy donors. We found: 1) the MDS-RS MSCs, estimated by colony forming unit-fibroblast (CFU-F), shared similar immunophenotype with normal MSCs (CD45-CD235a-CD31-CD44-, most of which were CD271+CD146+CD106+); 2) the frequency of CFU-Fs was significantly increased in the phenotypically defined MSCs of MDS-RS bone marrow compared to that of age-matched healthy controls (p=0.005); 3) multi-lineage differentiation assay revealed impaired osteogenic differentiation potential, but enhanced adipogenic differentiation potential of MDS-RS MSCs; 4) FACS analysis showed increased frequency of the adhesion receptor integrin α4 (ITGA4) in the CD44- MSCs from MDS-RS bone marrow (p=0.013); 5) Correspondingly, RNA-sequencing of the freshly isolated bone marrow MSCs and endothelial cells revealed altered gene expression profile of these cells in MDS-RS patients. Among those, ITGA4, ITGA11, ITGAE and ITGB1 are upregulated in the MDS-RS MSCs, indicating potential abnormal adhesive interaction of the MSCs with hematopoietic stem cells in the patients. In addition, the cell cycling gene KI67 is upregulated whereas cell cycle negative regulators, like CDKN1A and CDKN1C are downregulated in the MDS-RS MSCs, which is consistent with their increased CFU-F activity. Interestingly, we detected abnormal expression of hematopoietic growth factors such as downregulation of ANGPTL4 in the MDS-RS MSCs and upregulation of ANGPT1 in the MDS-RS endothelial cells. The functional relationship between the stromal cell alterations and the abnormal hematopoiesis as well as the underlying molecular mechanisms are currently under investigation.

Taken together, our data provide new evidence for phenotypic, functional and molecular alterations of bone marrow mesenchymal cells in MDS-RS patients. The molecular pathways mediating bone marrow niche alteration could be potential therapeutic targets for new treatment of MDS-RS.

Disclosures

No relevant conflicts of interest to declare.