bluuebunny/crossref_metadata_2025_split huggingface dataset:
Study of Renal Iron Overload by T2* MRI in a Large Cohort of Thalassemia Major Patients
Authors: Meloni Antonella , De Marchi Daniele , Positano Vincenzo , Giuffrida Gaetano , Armari Sabrina , Campisi Saveria , Furbetta Mario , Maggio Aurelio , Valeri Gianluca , Chiodi Elisabetta , Lombardi Massimo , Pepe Alessia 
Published: October 2019

Abstract
Abstract 5177

Background.
Renal dysfunction has been reported in adult subjects with thalassemia major (TM) since 1975. One of the main cause is the iron overload consequent to regular transfusions. Multiecho T2* MRI is a well-established technique for cardiac and hepatic iron overload assessment, but there very few report concerning the kidneys.
The aims of this study were to describe the T2* values of the kidneys in patients with TM, to investigate the correlation between renal and myocardial or hepatic siderosis and biventricular cardiac function.

Methods.
119 TM patients (58 men, 30. 7 ± 8. 2 years) enrolled in the Myocardial Iron Overload (MIOT) networks underwent MRI. For the measurement of iron overload, multiecho T2* sequences were used. The left ventricle was segmented into a 16-segments standardized model and the T2* value on each segment was calculated as well as the global value. In the liver, the T2* value was assessed in a single region of interest (ROI) in a homogeneous area of the parenchyma. For each kidney, T2* values were calculated in three different ROIs and were averaged to obtain a representative value for the kidney. The mean T2* value over the kidneys was also calculated. Cine images were obtained to quantify biventricular morphological and functional parameters in a standard way.

Results.
T2* values in the right kidney were significant lower than in the left kidney (40. 3±11. 9 ms vs 44. 1±12. 7 ms, P<0. 0001). The mean T2* value over the kidneys was 42. 2±11. 9 ms and 40 patients (33. 6%) had a pathological value (T2*<36 ms, lower limit of normal evaluated on 20 healthy subjects). The mean T2* value did not show a significant difference amongst men ad women (43. 2±11. 7 ms versus 41. 3±12. 1 ms, P=0. 378). The mean T2* values increased with age in a significant manner (r=0. 321, P<0. 0001). There was a significant negative correlation between serum ferritin levels and mean renal T2* values (r=-0. 446, P<0. 0001). Significant positive correlations of the mean T2* values were demonstrated for liver (r=0. 511, P<0. 0001) and global heart (r=0. 262, P=0. 004) T2* values (Figure 1). No correlation was found between renal iron overload and bi-ventricular function parameters.

Conclusions.
Systemic T2* differences between left and right kidneys were found, with significant lower values in the right one. Mean T2* value increased with age. We confirmed that kidney iron deposition was not very common in TM, but it was correlated with iron deposition in liver and heart.

Disclosures:
No relevant conflicts of interest to declare.
Genetic Heterogeneity of Alpha - and Beta - Globin Mutations in a German Multiethnic Population Group
Authors: Grosse Regine , Oyen Florian , Weber Belinda , Schneppenheim Reinhard 
Published: October 2019

Abstract
Abstract 5185
Thalassemia syndromes are among the most common hereditary diseases worldwide and are widespread throughout the Mediterranean Region, Africa, the Middle East, India, Burma, the Southeast Asia and Indonesia. Similarly, hemoglobinopathies are common in different ethnic groups and mixed syndromes of Thalassemia and hemoglobinopathies exist.
In this retrospective study we report the number and type of Thalassemia mutations and their combination with hemoglobinopathies detected mostly in individuals with a migration background now living in Germany.
DNA samples were analyzed by polymerase chain reaction (PCR) and direct DNA-sequencing and additionally by multiplex ligation-dependent amplification (MRC-HOLLAND MLPA®).
Mutations in one or both beta-globin chains were found in 77 patients. Most of these mutations were beta Thalassemia mutations, n=41 (Thalassemia minor n=33, Thalassemia major/intermedia n=7, homozygous Quin-Hai Hemoglobinopathy n=1).
In 34 samples we found the hemoglobin S single base mutation c. 20A>T p. E7V, either alone or in combination with a hemoglobin C mutation c. 19G>A p. E7K or beta Thalassemia mutation (HbAS n=14, HbSS n=12, HbSC n=6, HbSThal n=2).
In two patients we found only the hemoglobin C mutation by itself (HbAC n=1, HbCC n=1).
In 52 samples we found mutations on the alpha-globin chains, most often the 3. 7KB deletion (n=27), followed by the SEA (n=4), the 20. 5 KB deletion (n=4), the 4. 2 KB deletion (n=4), the Dutch deletion (n=3) and alpha triplication (n=3). Two of the remaining seven samples showed new alpha Thalassemia mutations which have not been described yet.
In an additional 34 DNA samples we found a combination of alpha- and beta-globin chain changes. In 4 of these samples we detected new mutations in the alpha-globin chains.
The prevalence of mutations in the alpha- and beta-globin chains varies greatly because of a complex ethnic structure of our patients. Changes of the alpha-globin chains could not be safely detected by hemoglobin-electrophoresis. These situations are very important for genetic counseling in a population in which consanguineous marriages are common. Furthermore, alpha-globin mutations are genetic modifiers for beta Thalassemia and sickle cell disease and will influence the phenotype of beta-globin chain mutations. Patients with a compound heterozygous mutation for beta Thalassemia will probably not be detected by hemoglobin-electrophoresis alone, as in milder forms and early childhood the hemoglobin is not severely decreased and hemoglobin electrophoresis may be misinterpreted for Thalassemia minor.
Our findings underline the heterogeneity of beta-globin and alpha-globin chain mutations and the importance of hematological and molecular analyses in the diagnosis and genetic counseling.

Disclosures:
No relevant conflicts of interest to declare.
Adenoviral E4 Gene Stimulates Secretion of PEDF That Maintains Long-Term Survival of Human Glomerulus-Derived Endothelial Cells
Authors: Nekhai Sergei , Kumari Namita , Obuhkov Yuri , Jerebtsova Marina 
Published: October 2019

Abstract
Abstract 5187
Renal glomerular endothelial cells are specialized cells with an important role in physiological filtration and glomerular disease. However, maintenance of human primary endothelial cells requires stimulation with serum and growth factors that often results in modification of the cells properties. Previously, expression of early adenovirus region E4 was shown to help maintaining long-term survival of human endothelial cells in serum free media without addition of growth factors. In the current study, we showed that media conditioned with human epithelial cells stably transfected with Ad E4 region supported survival of human glomerulus-derived endothelial cells in serum-free media (Fig. 1 A and B). Mass-spectrometry analysis of the conditioned media identified pigmental epithelium derived factor (PEDF) as a major component of the conditioned media. PEDF expression in 293-E4 cells was validated by RT- PCR (Fig. 1C), Western Blot and ELISA analysis (Fig. 1D). PEDF expression was detected in mouse glomeruli. Supplementation with recombinant PEDF supported survival of primary endothelial cells and the cells transformed with SV40 large T antigen in serum-free media, and extended the life-span of both cell cultures. PEDF did not inhibit FGF-2 stimulated growth and tubulogenesis of endothelial cells. Thus we demonstrated that adenoviral E4 region stimulated expression and secretion of PEDF by human renal epithelial cells that acted as a survival factor for glomerulus-derived endothelial cells.

Acknowledgments:
This work was supported NIH Research Grants SC1GM082325, R25 HL003679, 2G12RR003048, 8G12MD007597, K25GM097501 and 1P30HL107253. This study was also supported in part by National Kidney Foundation grant.

Disclosures:
No relevant conflicts of interest to declare.
BCR-ABL1 Kinase Inhibits DNA Glycosylases to Enhance Oxidative DNA Damage and Stimulate Genomic Instability
Authors: Slupianek Artur , Falinski Rafal , Znojek Pawel , Stoklosa Tomasz , Flis Sylwia , Doneddu Valentina , Synowiec Ewelina , Blasiak Janusz , Bellacosa Alfonso , Skorski Tomasz 
Published: October 2019

Abstract
Abstract 520
Tyrosine kinase inhibitors (TKIs) such as imatinib, dasatinib and nilotinib revolutionized the treatment of BCR-ABL1 kinase-positive chronic myeloid leukemia in chronic phase (CML-CP). Unfortunately, 15–25% of patients initially responding favorably to imatinib will develop acquired drug resistance, which in 40–90% of cases is caused by genomic instability resulting in the appearance of clones expressing TKI resistant BCR-ABL1 kinase mutants. We reported that CML-CP leukemia stem and progenitor cell populations accumulate high amounts of reactive oxygen species (ROS) resulting in excessive oxidative DNA damage such as oxidized DNA bases (8-oxoguanine and 5-hydroxycytosine→uracil) (Nieborowska-Skorska et al., Blood, 2012). Unfaithful and/or inefficient repair of these lesions generates TKI resistant point mutations in BCR-ABL1 kinase. Oxidative DNA lesions may be removed by base excision repair (BER) or, if not removed, will create mismatches, which are repaired by mismatch repair (MMR). Since we found that MMR is inhibited in CML-CP (Stoklosa et al., Cancer Res., 2008), the activity of BER is critical to prevent the accumulation of point mutations. Using an array of specific substrates and inhibitors/blocking antibodies we found that two major glycosylases, uracil-DNA glycosylase UNG2 and 8-oxoguanine glycosylase (OGG1) responsible for the excision of uracil (product of oxidation of cytosine) and 8-oxoguanine (8-oxoG) from DNA, respectively, were inhibited in BCR-ABL1 –transformed cell lines and CD34+ CML cells. The inhibitory effect was even more pronounced in CML blast phase (CML-BP) in comparison to CML-CP, it depended on BCR-ABL1 kinase activity and was not accompanied by deregulation of nuclear expression and/or chromatin association of these glycosylases. The effect was BCR-ABL1 kinase-specific because several other fusion tyrosine kinases such as TEL-ABL1, TEL-PDGFbetaR and NPM-ALK did not reduce UNG2 activity. Using UNG2-specific inhibitor UGI we found that UNG2 activity diminished the number of oxidized DNA bases detected by modified comet assay and prevented accumulation of point mutations in reporter gene Na+/K+ATPase, which encode resistance to ouabain. In conclusion, we hypothesize that inhibition of UNG2 and OGG1, accompanied by reduced MMR activity is responsible for accumulation of TKI-resistant BCR-ABL1 kinase point mutations and perhaps also other point mutations facilitating malignant progression of CML.

Disclosures:
No relevant conflicts of interest to declare.
Refinement in Patient Selection to Reduce Treatment-Related Mortality From Stem Cell Transplantation in Amyloidosis
Authors: Gertz Morie A , Lacy Martha Q , Dispenzieri Angela , Kumar Shaji K. , Dingli David , Leung Nelson , Hogan William J , Buadi Francis K , Hayman Suzanne R 
Published: October 2019

Abstract
Abstract 599

Purpose:
This study was undertaken to develop selection guidelines for transplant centers to determine eligibility for stem cell transplantation in patients with light chain amyloidosis.

Patients and Methods:
Patients with biopsy-confirmed immunoglobulin light chain amyloidosis who underwent stem cell transplantation between March 8, 1996, and December 31, 2011, were reviewed in 2 cohorts: those who underwent transplantation between March 8, 1996, and June 30, 2009, and those who underwent transplantation between July 1, 2009, and December 31, 2011 (table 1). A second comparison was undertaken among patients who died before posttransplant day 100 to determine features predictive of early death (table 2).

Results:
A total of 499 patients were identified, 410 in the earlier group and 89 in the later group. After July 1, 2009, significantly fewer transplant recipients had Mayo stage III cardiac involvement. Mortality before posttransplant day 100 was 10.5% (43/410) in the earlier group and 1.1% (1/89) in the later group. In the earlier group, one-quarter of transplant recipients with N-terminal pro-brain natriuretic peptide (NT-proBNP) levels higher than 5,000 pg/mL died by 10.3 months (fig 1, left). When the serum troponin T level was > 0.06 ng/mL, 25% died at 3.7 months (fig 2, right).

Conclusion:
The Mayo staging system is highly predictive for overall survival but not useful for selecting transplant recipients. Patients with serum troponin T levels higher than 0.06 ng/mL or NT-proBNP levels >5,000 pg/mL (not on dialysis) should not be considered acceptable candidates for stem cell transplantation because of unacceptable early mortality.

Disclosures:
No relevant conflicts of interest to declare.
MLL-Rearranged Leukemias Are Dependent On the Histone H2B Ubiquitin Ligase RNF20
Authors: Vakoc Christopher R. , Wang Eric , Kawaoka Shinpei 
Published: October 2019

Abstract
Abstract 649
Histone mono-ubiquitination is a major signaling mechanism that shapes the structure and function of chromatin. Hence, factors that regulate ubiquitin attachment to histones (E2 conjugating enzymes, E3 ligases, and de-ubiquitinating enzymes) often play fundamental roles in epigenetic biological processes, including the pathophysiology of human disease. Here, we explored whether the known regulators of histone ubiquitination/deubiquitination are linked to the pathogenesis of acute myeloid leukemia, a disease often associated with aberrant chromatin signaling. To this end, an shRNA screen was performed where all histone E2s, E3s, and deubiqitinating enzymes were individually suppressed and evaluated for their role in supporting growth of acute myeloid leukemias initiated the MLL-AF9 oncogene. This analysis identified the histone H2B ubiquitin ligase RNF20 (also known as BRE1A) as essential for MLL-AF9 leukemia growth. This finding was verified in a genetically engineered mouse leukemia model (driven by MLL-AF9 together with NrasG12D) under both in vitro and in vivo conditions. In addition, human leukemia cell lines harboring MLL-rearrangements were found to be uniformly dependent on RNF20 for their proliferation. Upon inhibition of RNF20 and elimination of global H2B mono-ubiquitination, leukemia cells arrest in the G1/G0 stage of the cell-cycle without evidence of myeloid maturation. RNA-SEQ analysis identified that a suite of direct transcriptional targets of MLL-AF9 were downregulated in expression upon RNF20 inhibition, including Hoxa9, Hoxa10, Meis1, and Mef2c. While H2B mono-ubiquitination has been shown previously to promote the catalytic function of H3K4 and H3K79 methyltransferases, we find that in leukemia cells inhibition of RNF20 eradicates global H2B mono-ubiquitination without influencing global H3K4me3 and H3K79me2/me3. However, we find instead that RNF20 is selectively required to maintain local levels of H3K79 methylation in the body of MLL-AF9 target genes, such as Hoxa9. Since MLL-AF9 is known to directly recruit the H3K79 methyltransferase DOT1L to transactivate its target genes, our findings suggest a mechanism whereby targeting RNF20 interferes with the transactivation function of MLL-AF9. Moreover, we found that shRNA-based suppression of RNF20 sensitizes leukemia cells to the growth-arrest induced by the BET bromodomain inhibitor JQ1, without influencing the sensitivity to cytotoxic agents daunorubicin and etoposide. Based on these collective findings, we propose that RNF20 represents a major chromatin-based signaling pathway necessary to maintain the MLL-rearranged subtype of acute myeloid leukemia.

Disclosures:
No relevant conflicts of interest to declare.
Carfilzomib, Cyclophosphamide and Dexamethasone (CCd) for Newly Diagnosed Multiple Myeloma (MM) Patients
Authors: Palumbo Antonio , Bringhen Sara , Villani Oreste , Siniscalchi Agostina , Russo Eleonora , Uccello Giuseppina , Cerrato Chiara , Gilestro Milena , Rossi Davide , Boccadoro Mario 
Published: October 2019

Abstract
Abstract 730

Background:
Carfilzomib is a novel, irreversible proteasome-inhibitor with a significant activity and favourable toxicity profile. Here we evaluated efficacy and safety of the combination carfilzomib-cyclophosphamide-dexamethasone in elderly (≥ 65 years) newly diagnosed MM patients.

Methods:
The Bryant and Day two-stage design was used to evaluate both efficacy and safety. Patients received oral cyclophosphamide (300 mg/m2 on days 1,8,15), oral dexamethasone (40 mg on days 1, 8, 15, 22) and iv carfilzomib (20 mg/m2 on days 1, 2, and 36 mg/m2 on days 8, 9, 15, 16, cycle 1; 36 mg/m2 on days 1, 2, 8, 9, 15, 16, cycles 2–9) every 28 days for 9 cycles, followed by maintenance with iv carfilzomib (36 mg/m2 on days 1, 2, 15, 16) every 28 days until progression or intolerance.

Results:
Thirty-four patients were enrolled: median age was 70 years, 21% of patients were older than 75 years, 24% had reduced creatinine clearance (<50 mL/min), 46% had ISS stage III, and 28% had unfavorable FISH [t(4;14) or t (14;16) or del17p]. Nineteen patients were evaluated for response and for safety after at least 4 cycles (table). All patients achieved at least partial response (PR), 74% at least very good partial response (VGPR), 42% at least complete response (CR)/ near-CR, including 10% stringent-CR. Responses were rapid with the median time to PR of 1 month and the median time to CR of 2 months. After a median follow-up of 7.5 months no patient has progressed or died.
Grade (G) 4 hematologic toxicities included neutropenia (1 patient, 5%). At least one G3-4 non-hematologic event was reported in 4 patients (21%): pneumonia and bronchitis (2 patients, 10%), atrial fibrillation (1 patient, 5%) and renal failure (1 patient, 5%). Only 1 patient (5%) developed G1 peripheral neuropathy. No patient discontinued treatment and 4 patients (21%) required carfilzomib dose reductions due to adverse events (G4 neutropenia, G3 pneumonia, G3 atrial fibrillation and G3 renal failure).

Conclusions:
Carfilzomib-cyclophosphamide-dexamethasone showed encouraging activity in patients with newly diagnosed MM. The combination was well tolerated with a frequency of at least one G3-4 non-hematologic adverse event of 21%. Results will be updated at the meeting.

Disclosures:
Palumbo: Onyx: Honoraria. Bringhen:Onyx: Honoraria.
A Calmodulin Interacting Protein Iqcg Is Required for Definitive Hematopoiesis in Zebrafish
Authors: Chen Li-Ting , Liang Wen-Xue , Chen Shuo , Chen Zhu , Chen Sai-Juan 
Published: October 2019

Abstract
Abstract 764
We previously identified a novel fusion gene NUP98-IQCG in a bi-phenotypic acute T-lymphoid/myeloid leukemia patient. However, the function of IQCG has never been demonstrated. By taking advantage of the zebrafish as a model organism, we investigated the role of IQCG in hematopoietic development and found that the definitive hematopoiesis was severely impaired in the iqcg deficient embryos. The hematopoietic stem cell (HSC) population as well as the number of multilineage differentiated hematopoietic cells was reduced. Moreover, the number of phosphorelated histone3 positive HSC was decreased, which indicated a compromised proliferation in iqcg deficient HSC.
Since IQCG protein contains an IQ calmodulin binding motif (IQ motif), we next examined if IQCG protein can interact with calmodulin. Co-immunoprecipitation assay results showed that both human and zebrafish IQCG proteins have the ability to precipitate calmodulin. Deletion of IQ motif disrupted this interaction. The isothermal titration calorimetry results indicated that the IQ motif of IQCG protein have a lower association constant with calcium-loaded calmodulin than apo-calmodulin. We next crystallized the complex of calmodulin and IQ motif of IQCG in two circumstances-with and without calcium. Determination of these two crystal structures indicated different interaction details between them. These results suggest that IQCG may be involved in calcium signaling in HSC.
Previous studies reported that calmodulin dependent protein kinase IV (CaMKIV) involves in HSC maintenance and survival. So we wondered if CaMKIV acts downstream of IQCG. Indeed, whole-mount immunofluorescence data showed that the level of phosphorelated CaMKIV markedly reduced in iqcg morphants compared with controls. Reactivation of CaMKIV by co-injection with constitutively activated CaMKIV (CACaMKIV) mRNA rescued the HSC number in iqcg deficient embryos. Furthermore, embryos injected with camkIV morpholino phenocopied the iqcg morphants.
In summary, we demonstrate an important role of the novel NUP98 partner gene IQCG in definitive hematopoietic development. IQCG may control the HSC proliferation through interacting with calmodulin and acting upstream of CaMKIV, which suggests a novel pathway in definitive hematopoietic development.

Disclosures:
No relevant conflicts of interest to declare.
RB Controls Size, Cellularity, and T Cell Output of the Mouse Thymus
Authors: Garfin Phillip M. , Viatour Patrick , Min Dullei , Bryson Jerrod , Weinberg Kenneth I. , Manley Nancy R. , Sage Julien 
Published: October 2019

Abstract
Abstract 835
The establishment of the thymic microenvironment early in life is crucial for the production functional T cells. Conversely, thymic involution results in a decreased T cell output. Thymic involution has important health implications especially following bone marrow transplant. Our objective is to determine molecular and cellular mechanisms that will allow for regeneration of involuted thymic tissue, restore production of naïve T cells, and improve immune function while improving our understanding of immunobiology. In this pursuit, we have focused on the Retinoblastoma family of tumor suppressor proteins. The main function of the RB pathway is to restrict passage through the G1/S transition of the cell cycle. RB and its two family members, p107 and p130, mediate the action of a broad range of cellular signals to control the proliferation, survival, and differentiation status of a large number of mammalian cell types. We found that inactivation of the RB pathway in the thymus by early deletion of RB family genes prevents thymic involution, promotes expansion of functional thymic epithelial cells (TECs), and increases thymic T cell output. Moreover, we have identified a direct regulatory relationship between RB and the Foxn1 transcription factor Via E2F transcription factors, where RB/E2F complexes directly repress the Foxn1 promoter, thereby promoting involution. Thus, the RB family is a critical mediator of extra- and intra-cellular signals to regulate thymic epithelial cells and thymus function, and decreasing RB pathway function may promote regeneration of the involuted thymus and restoration of naïve T cell production in patients.

Disclosures:
No relevant conflicts of interest to declare.