bluuebunny/crossref_metadata_2025 huggingface dataset:
Dasatinib Cerebrospinal Fluid Concentration and Plasma Pharmacokinetics: Potential for Central Nervous System Prophylaxis In Philadelphia Chromosome-Positive Leukemia
Authors: Takahashi Naoto , Miura Masatomo , Scott Stuart , Saitoh Hirobumi , Motegi Mutsuhito , Fujishima Naohito , Kameoka Yoshihiro , Tagawa Hiroyuki , Hirokawa Makoto , Sawada Kenichi
Published: October 2019

Abstract 1807

Dasatinib (DA) is approved for use in imatinib-resistant or intolerant chronic myeloid leukemia (CML)/Philadelphia-positive acute lymphoid leukemia (Ph+ALL) and may also be useful for central nervous system (CNS) leukemia accompanied with CML/Ph+ALL; however, little is known about the relationship between DA pharmacokinetics and its ability to penetrate the blood-brain barrier. Consequently, we measured DA plasma and cerebrospinal fluid (CSF) levels by high-performance liquid chromatography in 20 samples obtained from 11 DA-treated patients (seven Ph+ALL and four lymphoid crisis CML). DA was detected in 10 CSF samples from five patients who were treated with 100 mg QD of DA (CSF C4h of detectable group; 3.526±2.604 ng/mL, 1.11–7.95 ng/mL), which was above the IC50 level for wild type BCR-ABL positive leukemia cells in vitro (0.8 nM = 0.39 ng/mL). However, DA was not detected in 10 CSF samples from 7 patients (CSF C4h of non-detectable group; <1.0 ng/mL). The concentration ratio of CSF to plasma was 3.90% (0.42-12.23%), which approached previously reported ratios for imatinib. There were significant differences in the AUC0-4 and the plasma C4h between the CSF detectable (D) and non-detectable (ND) patients (AUC0-4: 268.29±92.452 vs. 90.83±76.45, P=0.00019 by Student t-test, Figure 1; plasma C4h: 126.15±62.58 vs. 47.41±50.935, P=0.00637 by Student t-test). Moreover, there were significant correlations between CSF C4h and AUC0-4 (P<0.01, Figure 2) and between CSF C4h and plasma C2h (P<0.001), together suggesting that penetration of DA into the CSF may depend on DA plasma concentration. To investigate any influence of pharmacogenetic variation on CSF penetration, single nucleotide polymorphisms in genes involved in DA pharmacokinetics and transport (ABCB1, ABCG2, SLC22A1, SLC22A3, and CYP3A4/5) were interrogated; however, no significant correlation between CSF levels and genotype were observed. DA has a 325 fold greater potency than imatinib for inhibiting BCR-ABL tyrosine kinase, which undoubtedly influences the efficacy of DA for Philadelphia-chromosome positive CNS leukemia; however, our data suggest that clinical DA blood level monitoring may help estimate the penetration of DA to the CSF.

Disclosures:

No relevant conflicts of interest to declare.
DNA damage-induced Leukaemic Stem and Progenitor Cell Kill with Mylotarg and Tipifarnib/Zarnestra Drug Combination In Acute Myeloid Leukemia (AML)
Authors: Jawad Mays , Yu Ning , Russell Nigel H. , Pallis Monica
Published: October 2019

Abstract 1824

Leukaemic stem and progenitor cells (LSPC) are relatively resistant to current chemotherapy and considered to contribute to disease progression and relapse in Acute Myeloid Leukemia (AML) patients following initial treatment. We have developed a model to study the in vitro targeting of AML LSPC by novel agents which have the potential to target LSPC whilst in a resistant niche microenvironment. Using this model, we showed that under ‘niche-like’ conditions CD34+CD38-CD123+ LSPC are less sensitive to cytarabine and to the Flt3 inhibitor AG1296. In contrast, LSPC when grown under niche support are extremely sensitive to Mylotarg (Gemtuzumab Ozogamicin), one of a few successful chemotherapeutic agents that is currently used in the clinic. We are now proposing that the combination of Mylotarg (10ng/ml) and the farnesyltransferase inhibitor, Tipifarnib/Zarnestra (5microM) is successful at further targeting the LSPC subset resulting in 70% LSPC kill (n=29 AML primary cells) after 48hr in vitro treatment compared to 31% and 12% LSPC kill in Mylotarg and Tipifarnib treated cells, respectively (p<0.0001). Moreover, stromal MS5 co-culture experiments (used to depict drug-induced cell survival in a niche-like setting) with the Tipifarnib/Mylotarg combination showed enhanced cytotoxicity even in such a resistant setting. Because activation of alternative signaling pathways maybe one mechanism that cancer cells use to evade drug-induced cell kill, we used a protein profiler assay to depict the expression of 46 known human phosphokinase proteins when treated with single and drug combination. Using the CD34+CD38- TF1A cell line, 34/46 (74%) of phosphokinase proteins that are essential for functional survival pathways (e.g. p38MAPK, ERK, JNK, AKT, STAT1/2/4/5, GSK3beta, beta-catenin, p53 etc.) were maximally reduced by the drug combination compared to housekeeping phosphoproteins, 12/46 (26%) proteins were unaffected by the drug combination and only 1/46 (the cell cycle checkpoint regulator and putative tumor suppressor Chk2) increased maximally with the drug combination. We next show that the drug combination's success is indeed characterized by the activation of DNA damage response (increase in H2AX foci by immunoflourescence), an induction of phosphoATM (by flow cytometry) leading to the activation of Chk2 and G2/M cell cycle arrest (using BrdU assay). Finally, we demonstrate that Tipifarnib may be acting as a potent P-glycoprotein inhibitor by allowing the retention of Mylotarg in cells that would otherwise efflux the drug out of the cell. In conclusion, the in vitro data presented here clearly shows why we think the Tipifarnib/Mylotarg combination can be successful in the treatment of AML and warrants further investigation to confirm its efficacy in in vivo models.

Disclosures:

No relevant conflicts of interest to declare.
Temozolomide In Acute Myeloid Leukemia: A MGMT Promoter Methylation Status–Based Treatment Stratification
Authors: Medeiros Bruno C. , Kohrt Holbrook E , Rajwanshi Richa , Gotlib Jason , Coutre Steven , Liedtke Michaela , Berube Caroline , Zhang Melody , Arber Daniel A. , Zehnder James L.
Published: October 2019

Abstract 3313

Introduction:

Temozolomide sensitivity is determined by the methylation status of the MGMT promoter. As most AML patients have an unmethylated MGMT promoter, response rates to temozolomide have been very disappointing in the past. We designed this exploratory study to determine whether temozolomide therapy can be tailored according to the MGMT promoter status and to test the hypothesis that protracted, low-dose temozolomide can “prime” leukemia blasts to conventional doses of temozolomide in patients with unmethylated MGMT promoter.

Patients and Methods:

Elderly patients (>60 years of age) with AML and high-risk features (relapsed/refractory, secondary AML or de novo with intermediate or unfavorable cytogenetics) were stratified according to MGMT promoter methylation status. Patients with methylated MGMT promoters received temozolomide 200mg/m2 orally for 7 days, while patients with unmethylated promoters received temozolomide 100mg/m2 orally for 14 days followed by temozolomide 200mg/m2 orally for 7 days.

Results:

36 patients (median age, 75 years) were treated with temozolomide and 31 (86%) were found to have an unmethylated MGMT promoter. Overall response rate for the entire cohort was 36% (CR – 22%, CRp – 8%, LFS- 6%). The median duration of response and median overall survival among responding patients were 29 weeks and 35 weeks, respectively. No differences in outcomes (CR rate, OS rate and duration of remission) were noted among the 2 stratification groups (Table-1) Induction deaths (within 42 days of treatment initiation) occurred in 25% (9/36) of patients and were mostly caused by disease progression. Patients with low HCT-CI scores (≤ 2) had higher ORR 43% vs.0%, fewer induction deaths (17% vs. 67%, respectively) and longer median OS (94 days vs. 25.5 days). Hematological toxicities were the most commonly observed adverse events and difficult to distinguish from disease-related cytopenias.

Conclusion:

Temozolomide therapy, stratified according to MGMT promoter methylation status, demonstrated clinical activity in elderly patients with AML and high-risk features. Protracted low-dose temozolomide may reverse the temozolomide-resistant phenotype in patients with unmethylated MGMT promoters. Future studies in better-defined populations or in combination are warranted. This trial was registered at ClinicalTrials.gov as NCT00611247.

Disclosures:

Medeiros: Schering-Plough: Research Funding. Off Label Use: Temozolomide for elderly AML.
Comparasion Between Nilotinib and Dasatinib as Second-Line Therapy for Patients with Chronic Myeloid Leukemia: A Single Center Retrospective Study.
Authors: Lobo Clarisse , Boquimpani Carla , Madeira Tania Silva , Wendling Patricia , Maximo Claudia , Lima Carlos Bernardo , Atta Elias , DE SOUZA Cravo Renata
Published: October 2019

Abstract 3440

Nilotinib and dasatinib are second-generation tyrosine kinase inhibitors (TKI) used in patients with chronic myeloid leukemia (CML) resistant or intolerant to imatinib. There are no randomized clinical trials comparing these drugs in this context. The aim of this study was to compare, retrospectively, the hematological, cytogenetic and molecular response in patients submitted to these second-generation TKI at Hemorio, a public brazilian institution. A total of 114 patients were analyzed, 63 received nilotinib and 51 dasatinib as second-line therapy (55.3% and 44.7%, respectively). The following variables were equally distributed between these two groups (nilotinib vs. dasatinib, respectively): male sex (54% vs. 60.8%, p=0.46), median age at diagnosis (46 vs. 45 years, p=0.76), median time in months using imatinib before the switch (45.2 vs. 44.1, p=0.96), resistance to imatinib (98.4% vs. 98%, p=0.88), presence of the mutation T315I (3.2% vs. 3.9%, p=0.09), patients in chronic phase before the switch (85.7% vs. 86.3%, p=0.93). Use of another second generation TKI, as a third-line therapy, was necessary in 30 out of the 114 patients analyzed (26.1%) because of lack of response. This modification was slightly more frequent in the group initially submitted to nilotinib (31.7% vs. 19.6%, p=0.21). Patients who used a third-line therapy were excluded from response and survival analyzes. Response rates after the second-generation TKI were similar between these two groups (nilotinib vs. dasatinib): complete hematological response until three months (77.8% vs. 87.3%, p=0.24), complete cytogenetic response until six months (21.6% vs. 22.2%, p=0.95) and 12 months (32.4% vs. 33.3%, p=0.94) and major molecular response reached before 12 months (32.7% vs. 21.6%, p=0.25). Two-year overall survival (OS) and progression free-survival (PFS) were similar between these two groups (nilotinib vs. dasatinib, respectively): 92.2% vs. 87.8% (p=0.38) for OS and 87.8% vs. 83.7% (p=0.14) for PFS. Although not statistically significant, two-year OS was inferior in the group of patients who needed a third-line therapy (70.5% vs. 95.6%, p=0.70). Our results suggest that the response and survival rates are similar between nilotinib and dasatinib as second-line therapy for patients with imatinib resistant or intolerant CML. Also, they suggest an inferior prognosis for patients who need a third-line therapy. In this way, the choice between these two TKI for second-line therapy should be guided by the clinical characteristics and the mutation status of the patient.

Disclosures:

Lobo: NOVARTIS: Research Funding.
C-Myc Inhibition Targets the Leukemia-Initiating Cells in a Tal1/Lmo2 mouse Model of T-ALL
Authors: Tesell Jessica , Kelliher Michelle , Hermance Nicole , Roderick Justine E. , Bradner James E
Published: October 2019

Abstract 1880

Thymic expression of the Tal1 and Lmo2 oncogenes in mice results in rapid development of T-ALL, and similar to T-ALL patients, over half the leukemic mice develop spontaneous mutations in Notch1. We have previously shown that Notch1 inhibition reduces and in some cases eliminates leukemia-initiating cells (L-ICs), however the use of anti-Notch1 therapies in the clinic has been limited due to on-target effects of Notch inhibition in the intestine. Due to the toxicities associated with Notch inhibition in vivo, we hypothesized that it may be more effective to inhibit c-Myc, a critical Notch1-regulated gene in T-ALL. We demonstrate that silencing of c-Myc in primary mouse T-ALL cells significantly prolongs survival by reducing L-IC frequency and self-renewal. A novel bromodomain inhibitor, JQ1, has been developed and shown to inhibit c-Myc in multiple hematopoietic malignancies, but its effects on LSCs or L-ICs remain unclear. Our preliminary studies reveal that JQ1 treatment prolongs survival in our mouse T-ALL model, at least in part, by reducing c-Myc mRNA and protein levels. Whether JQ1 has anti-L-IC activity in mouse T-ALL models and relapsed pediatric T-ALL patient samples will be discussed.

Disclosures:

No relevant conflicts of interest to declare.
Influence of recipient's Age On the Outcome of HLA-Matched Sibling Transplants in Adult Patients with Severe Aplastic Anemia Who Conditioned with Fludarabine-Based Regimen
Authors: Shin SeungHwan , Yoon JaeHo , Yahng SeungAh , Lee SungEun , Cho ByungSik , Eom KiSung , Kim YooJin , Lee Seok , Min ChangKi , Kim Heeje , Cho SeokGoo , Kim DongWook , Min WooSung , Park ChongWon , Lee JongWook
Published: October 2019

Abstract 1921

Background:

There is a controversy concerning upper age limit of recipient in allogeneic stem cell transplantation (allo-SCT) of severe aplastic anemia (SAA) because older patients have an increased risk of transplant-related mortality (TRM). Recently, a less toxic regimen comprising reduced dose of cyclophosphamide (CY), fludarabine (Flu), and ATG (Flu/CY/ATG) was introduced for conditioning the patients with SAA scheduled for HLA-identical sibling allo-SCT. We analyzed the outcome of these patients after conditioning with Flu/CY/ATG regimen according to recipient's age.

Method:

We analyzed 82 consecutive adult patients transplanted from March 2002 to June 2011, followed conditioning with CY (50 mg/kg/day × 2 days), Flu (30 mg/m2/day × 6 days), and ATG (2.5 mg/kg/day × 4 days).

Results:

The median age of patients was 39 years (range, 13–58 years). 29 (35.4%) patients were very SAA at the time of allo-SCT. Stem cell sources were bone marrow (BM, n=59), peripheral blood stem cell (PBSC, n=12), and BM+PBSC (CD34+ cell selection, n=11). The conditioning regimen was well tolerated, with low TRM (4.9%). Neutrophil engraftment was observed in all patients; the median time to engraftment of neutrophils and platelets was 12 and 17 days, respectively. The 100-day cumulative incidence (CI) of acute graft-versus-host disease (GVHD, grade °Ã2) was 3.7% and 2-year CI of chronic GVHD was 4.4%. With a median follow-up of 3 years, overall survival (OS) and failure-free survival was 93% and 83%, respectively. Seven patients (8.5%) developed delayed graft failure and 6 patients of them have successful durable engraftment after second allo-SCT without TRM. Seven patients died of veno-occlusive disease (n=1), cytomegalovirus pneumonia (n=2), sepsis (n=2), clonal evolution after allo-SCT (n=2, 1 acute myeloid leukemia and 1 myelodysplastic syndrome), respectively. 3-year probability of OS by age group were 88% at less than 20 years old (n=8), 97% at 20–39 (n=34), 92% at 40–49 (n=26), 86% at 50–59 (n=14). There is no significant difference in OS according to age group (p=0.426).

Conclusion:

These data indicate that the Flu/CY/ATG conditioning regimen is well tolerable with low TRM and CI of GVHD in all age group. Similar OS according to 4-divided age group suggest that patient's age more than 50 years old might be possible candidate for allo-SCT in SAA with Flu/CY/ATG conditioning regimen.

Disclosures:

No relevant conflicts of interest to declare.
Unmanipulated Graft Transplantation From Family Haploidentical Donors: A Survey On 183 Adult Patients with Acute Leukemias On Behalf of the Acute Leukemia Working Party of the European Group for Blood and Marrow Transplantation (EBMT)
Authors: Ciceri Fabio , Labopin Myriam , Di Bartolomeo Paolo , Arcese William , Santarone Stella , De Angelis Gottardo , Peccatori Jacopo , Mohty Mohamad
Published: October 2019

Abstract 1987

Background.

Allogeneic stem cell transplantation from an haploidentical family donor has been recently developed through unmanipulated graft platforms.

Methods.

We collected in 57 EBMT centers 183 haploidentical transplants (haploSCT) from unmanipulated peripheral blood (PB) and bone marrow (BM) graft in adults patients (pts) with de novo acute myeloid leukemia (AML) and acute lymphoblastic leukemia (ALL) reported to EBMT registry from 2007 to 2010 and analysed the outcome according to the known risk factors.

Results.

Overall, 120 AML patients underwent transplantation in CR1 (39), CR2 (22) or in advanced disease (54). Overall, 63 ALL pts underwent transplantation in CR1 (26), CR2 (14) or in advanced disease (22). Median age was 42y (18–75). Graft composition was based on non ex-vivo T-cell depleted BM in 52 (28%) or PB in 131 (72%). Conditioning regimen was myeloablative (MAC) in 95 and reduced-intensity (RIC) in 88 pts. Primary engraftment was documented in 160 pts (88%), with ANC 0.5 ×109/L in a median of 19 days (BM) and 16 days (PB). Graft-versus-Host Disease (GvHD) prophylaxis was based on Anti-thymocyte globulins (ATG) + cyclosporine-methotrexate (24%) or cyclosporine-mycophenolate (18%), rapamycine-mycophenolate (25%), tacrolimus-mycophenolate (15%), others (7%). The cumulative incidence of acute GvHD >=II was 26%. With a median follow-up of 13 months (1–49), the estimated leukemia-free survival (LFS) at 1 years i was 53 +/−7%, 36 +/−8% and 22 +/−5% for CR1, CR2 and advanced patients respectively. In multivariate analysis for competing risks, relapse incidence (RI) at 1 y was 23 +/−5%, 20 +/−7%, 48 +/−6% for CR1, CR2 and advanced. The non-relapse mortality (NRM) at 1 y was 23 +/−6%, 43 +/−5%, 29 +/−6% for CR1, CR2 and advanced patients. On multivariate analysis, the only factor relevant for LFS was disease status at transplant.

Conclusion.

EBMT registry analysis of haploidentical transplantation from unmanipulated donor graft confirms relevant LFS for patients with high risk acute leukemia. These results suggest that this procedure should be considered prospectively int treatment algorithm for high risk adults patients in early stages.

Disclosures:

No relevant conflicts of interest to declare.
Impact of CD3/T Regs Ratio in Donor Graft On Survival Rates in Allogeneic Peripheral Blood Stem Cell Transplantation
Authors: Delia Mario , Pastore Domenico , Mestice Anna , Carluccio Paola , Perrone Tommasina , Gaudio Francesco , Curci Paola , Ricco Alessandra , Rossi Antonella Russo , Giordano Annamaria , Carluccio Vera , Di Tardo Ilaria , Angarano Rosa , Longo Maria Chiara , Specchia Giorgina
Published: October 2019

Abstract 2030

Background:

The therapeutic efficacy of allogeneic stem cell transplantation (alloSCT) for hematological malignancies relies largely on the graft versus leukemia (GvL) effect exerted by the donor CD3 cells, but an uncontrolled graft-versus-host-disease (GvHD) bears a risk of complications. On the other hand, T regs cells (CD4+CD25high Foxp3+) are believed to maintain tolerance and to inhibit GvHD after alloSCT; also, the Foxp3 gene encodes a transcription factor that is a key for thymic development, so T regs cells could preserve an optimal microenviroment for the reconstitution of functional immunity after alloSCT. Moreover, when looking at post allotransplant patients' outcomes, while it is largely known the impact of acute GVHD (triggered by CD3 donor T cells) on survival, there is no evidence that donor graft CD3/T regs ratio may determine an effect in terms of OS, NRM and relapse free survival rates so far.

Patients and Methods:

In this study we analyzed the graft CD3+/Tregs ratio (gCD3/Tregs R) and determined its impact on acute GVHD (aGVHD), immunological recovery and survival rates (OS, NRM and Relapse) after myeloablative alloPBSCT. We analyzed 74 consecutive patients transplanted with unmanipulated peripheral blood stem cells from an HLA identical related donor (n=48) or an HLA identical unrelated donor (n=26); diagnoses were acute myeloid leukaemia (n=62), acute lymphoblastic leukaemia (n=13). The median CD3+ and Tregs dose administered was 238 (range (r): 67–550) and 12,5×106̂/Kg (r: 2–21), respectively; the median gCD3/Tregs R was 19 (r: 8–250). Patients were subdivided into a high gCD3/Tregs R (>=36) group (HR group n= 30) and a low gCD3/Tregs R (<36) (LR group n=44).

Results:

The incidence of aGVHD (grade II-IV) in the low gCD3/Tregs R (LR) group was lower than in the high gCD3/Tregs R (HR) group (4/30 or 13% vs 36/44 or 82%, p<.001). The OS, NRM and relapse rate at 3 years was 54,29 and 34%, respectively. Comparing LR with HR group a statistically significant difference is demonstrated for OS and NRM rates (65 vs 31%,p<.004; 3 vs 71%, p<.001), respectively, but not for the R one (35 vs 30%, p=ns). Comparing aGVHD+ with aGVHD- group OS, NRM and relapse were always statistically significant different (39 vs 65%,p<.005; 61 vs 7%,p<.001; 9 vs 53%,p<.002).

Conclusions:

Taken together, our data may suggest that Tregs content is able to mediate protective effects against aGVHD, while preserving GvL effects as demonstrated by relapse rate comparison between H and LR groups. However, larger studies are needed to understand the real contribution of gCD3/Tregs R on survival rates.

Disclosures:

No relevant conflicts of interest to declare.
Three Novel EPAS1/HIF2A Somatic and Germline mutations Associated with Polycythemia and Pheochromocytoma/Paraganglioma.
Authors: V Felipe R Lorenzo , Zhuang Zhengping , Yang Chunzhang , Fui Mark Ng Tang , Vankayalapati Hariprasad , Pacak Karel , Prchal Josef T.
Published: October 2019

Abstract 2080

Congenital polycythemias have diverse etiologies, including mutations in the hypoxia sensing pathway leading to increased levels of hypoxia inducible factors (HIF) and erythropoietin. These include mutations of negative regulators of HIFs, such as germline von Hippel-Lindau (VHL) gene heterozygous mutations (Chuvash polycythemia), HIF-prolyl hydroxylase 2 (PHD2) gene mutations, and gain-of-function mutations of HIF-2-alpha (HIF2A) (exon 12). Mutation of the PHD2 gene in one family was associated with polycythemia and recurrent pheochromocytoma/paraganglioma (PHEO/PGL) a neuroendocrine tumor commonly found in so called VHL tumor predisposition syndrome. Over the past two decades, we have studied six unrelated patients with sporadic congenital polycythemia who subsequently developed PHEO/PGLs without till now discernible molecular basis. We now report on these three patients, two with different tumor somatic codon A530 HIF2A mutations in exon 12 and one with germline mutation F374Y inherited from his mother, in a unique domain (exon 9) of HIF2A, all three cases later developed multiple recurrent neuroendocrine tumors and were subjects of our study.

In search of species homology, codons F374 and A530 of the HIF2A are highly conserved among man, chimpanzee, mouse, horse, cattle, chicken and zebrafish, suggesting this serves an important function for the gene. These mutations were identified in the vicinity of the primary hydroxylation site (exon 12) and novel domain (exon 9) of the HIF2A protein which affects VHL protein binding, Functional studies of the HIF2A mutants shown that three HIF2A variants have increased half-life consistent with gain-of-function of the HIF2A due to disruption in the VHL binding to these mutant residues impairing ubiquitination and proteasomal degradation. This results in increased transcription of genes downstream of HIF2A including erythropoietin. Further examination did not reveal any evidence of loss-of-heterozygosity, nor an additional mutation of HIF2A, or other HIF-pathway genes in their tumor tissues. The fact that two patients with polycythemia and PHEO/PGL had somatic and one germline HIF2A mutations, albeit at different locations, underscore the PHEO/PGL promoting potential of gain-of-function mutations of HIF2A that alone, either as somatic or germline mutations can contribute to, but are not sufficient for PHEO/PGL development but is sufficient for inducing polycythemic phenotype.

Disclosures:

No relevant conflicts of interest to declare.
A Hepcidin Inhibitor Mobilizes Iron for Incorporation Into Red Blood Cells in an Adenine-Induced Chronic Kidney Disease Model in Rats.
Authors: Vaja Valentina , Sun Chia Chi , Cappellini Maria D , Lin Herbert Yih-Fuu , Babitt Jodie L
Published: October 2019

Abstract 2082

Background:

Anemia is prevalent in patients with chronic kidney disease (CKD). A key feature of the anemia of CKD is limited iron availability for efficient erythropoiesis despite adequate body iron stores. It is now well established that excess levels of the iron regulatory hormone hepcidin in CKD are responsible for decreasing expression of the iron exporter ferroportin, thereby blocking iron absorption from the diet and iron release from body stores. Adenine treatment in rats has been proposed as an animal model of anemia of CKD with high hepcidin levels that mirrors the condition in patients.

Methods:

We developed a modified adenine-induced kidney disease model of anemia in rats by giving a diet supplemented with 0.75% adenine for 3 weeks followed by a normal diet for 3 weeks. We then tested whether the small molecule bone morphogenetic protein (BMP) inhibitor LDN-193189, which has previously been shown to lower hepcidin levels, was able to mobilize iron into the plasma and improve iron-restricted erythropoiesis in adenine-treated rats.

Results:

The modified adenine model had a higher survival rate than previously reported models, while maintaining irreversible renal failure and anemia. Adenine-treated rats had increased hepatic hepcidin mRNA, decreased serum iron, increased spleen iron content, low hemoglobin, and inappropriately low EPO levels relative to the degree of anemia. LDN-193189 lowered hepatic hepcidin mRNA and mobilized stored iron into plasma in adenine-treated rats. Moreover, the iron was efficiently incorporated into hemoglobin in reticulocytes. However, LDN-193189 alone did not prevent anemia progression in our model.

Conclusions:

Lowering hepcidin improved iron availability, but did not improve anemia in an adenine-induced kidney disease model in rats. Co-administration of hepcidin lowering agents with erythropoiesis stimulating agents (ESAs) may be useful as a combination therapy to correct iron balance and thereby reduce the ESA dose needed to achieve target hemoglobin levels.

Disclosures:

Sun: FerruMax Pharmaceuticals Inc: Consultancy. Lin:Ferrumax Pharmaceuticals: Consultancy, Equity Ownership, Patents & Royalties. Babitt:Ferrumax Pharmaceuticals, Inc: Equity Ownership.
Elevated Hematocrit Concentration and the Risk of Mortality and Vascular Events in Patients Undergoing Major Surgery.
Authors: Musallam Khaled M , Porter John B , Soweid Assaad , Hoballah Jamal J , Sfeir Pierre M , Khalife Mohamed , Zeineldine Salah , Tamim Hani M , Richards Toby , Lotta Luca Andrea , Peyvandi Flora , Jamali Faek R
Published: October 2019

Abstract 2088

Background:

Preoperative anemia is associated with adverse outcomes after major surgery. This study evaluates the effect of elevated hematocrit concentration on 30-day postoperative mortality and vascular events in patients undergoing major surgery.

Methods:

We conducted a cohort study using the American College of Surgeons National Surgical Quality Improvement Program database. Thirty-day mortality and vascular events, demographic, and perioperative risk factors were obtained for 197,469 adult patients undergoing major surgery in nonveteran's administration hospitals across the US, Canada, Lebanon, and the UAE during 2008 and 2009. We assessed the adjusted effect of elevated (>0.50) compared to normal preoperative hematocrit concentration (≥0.41–0.50, American Medical Association reference-range) on postoperative outcomes. Separate sex-specific analysis using hematocrit concentration thresholds commonly used in the diagnosis and management of patients with apparent or absolute erythrocytosis was also done.

Results:

A total of 3,961 patients (2.0%) had elevated hematocrit concentration preoperatively. After adjustment, postoperative mortality at 30 days was higher in patients with elevated hematocrit concentration than in those without (odds ratio [OR]: 2.23, 95% CI: 1.77–2.80). 30-day deep vein thrombosis (OR: 1.95, 95% CI: 1.44–2.64) and pulmonary embolism (OR: 1.79, 95% CI: 1.17–2.73), but not myocardial infarction or cerebrovascular events, were also higher in patients with elevated hematocrit concentration than in those without. Similar evaluation of various clinically relevant hematocrit concentrations revealed the following: an effect on mortality was noted beyond the thresholds of 0.48 in women and 0.52 in men, with the effect estimates becoming considerably high for values >0.54. Values between 0.41–0.45 were not associated with increased odds mortality. Similar observations were noted for deep vein thrombosis, although with higher variation and uncertainty especially in women; while the effects on pulmonary embolism were restricted to men.

Conclusion:

Elevated hematocrit concentration is associated with an increased risk of 30-day mortality and venous thrombosis following major surgery. Further investigation of the impact of elevated hematocrit concentration and its reduction on surgical outcomes is warranted.

Disclosures:

No relevant conflicts of interest to declare.
Homozygous Expression of a Novel Senegalese-Type Partial Deletion of the Beta and Delta Genes Causes a Delta0 Beta+ Thalassemia.
Authors: Sabio Hernan , Dixon Natalia , Kutlar Ferdane , Patel Niren , Zhang Hanfang , Zhuang Lina , Kutlar Abdullah
Published: October 2019

Abstract 2128

Clinical phenotype in β-thalassemia syndromes is determined by the degree of chain imbalance. An increase in γ-globin production will compensate for the absent or deficient β-globin synthesis and will result in the amelioration of the chain imbalance, and hence an improvement in clinical features. The known genotypes of δβ-thalassemia are associated with an increase in Hb F production, which results in the amelioration of the clinical presentation. Most δβ-thalassemias result from deletions that remove the δ- and β-globin genes, (δβ)0 with a compensatory increase in γ-globin (Hb F) expression. We report an unusual case of homozygous δ0β+ thalassemia that provides interesting insights into increased γ-globin expression and the regulation of β-globin gene expression. An 8-year old boy of African ancestry presented with lifelong jaundice and pallor. He also experienced episodes of worsening symptoms. He exhibited frontal bossing, pale mucosa, scleral icterus, and moderate splenomegaly. He was known to have G6PD deficiency and was suspected of having additional erythrocyte pathology. The CBC revealed a Hb of 8.7, Hct 26.4, MCV 64.7, WBC 10,700, platelets 283,000, reticulocytes 2.2%, and total bilirubin 5.3. Hemoglobin analysis by HPLC and IEF revealed HbA 13.4%, Hb F 86.6%, and no additional components. Alpha thalassemia −3.7kb deletion was not detected. Globin chain analysis revealed α, β, Gγ and AγI chains. DNA analysis revealed a novel Senegalese-type deletion of the beta and delta genes, resulting in a delta0 beta+ thalassemia. The subject's parents who were both from the same small village in Niger had normal hematology values. Their hemoglobin analyses revealed Hb A 94. 8%, Hb A2 2.0%, Hb F 3.2% and Hb A 93.5%, Hb A2 2.1%, Hb F 4.5% in the father and mother, respectively. They were both heterozygous for the delta-beta deletion identified in their son. DNA analysis revealed a breakpoint in the delta gene at nucleotides 54755–54760 and a breakpoint in the beta gene at nucleotides 62153– 62158 [GenBank Ref ID: HUMHBB] with a 5 nucleotide “CAACA” bp region overlapping area. The subject, who is homozygous for the identified deletions, has a clinical phenotype of thalassemia intermedia. He has not yet required red cell transfusions. This is the first instance of a Senegalese-type deletion occurring in the homozygous state. The genotype provides insights into regulation of globin gene expression. While the ∼7 Kb deletion in the δβ-intergenic region may be responsible for the increased expression of the γ-globin gene similar to Hb Lepore deletions, the continued low level expression of the β-globin gene is most probably the result of the juxtaposition of the inefficient δ-globin promoter brought in the vicinity of the β-globin gene.

Disclosures:

No relevant conflicts of interest to declare.
Platelet ERp57 Is Required for Hemostasis and Thrombosis.
Authors: Wang Lu , Wu Yi , Zhou Junsong , Ahmad Syed S. , Mutus Bulent , Essex David W
Published: October 2019

Abstract 2168

Several members of the protein disulfide isomerase family of enzymes are important in platelet function and in thrombosis. Platelet protein disulfide isomerase (PDI) has been shown to have an important role in platelet function but is reported to not be required for thrombus formation in vivo. A novel platelet PDI called ERp57 mediates platelet aggregation but its role in thrombus formation is unknown. To determine the specific role of platelet-derived ERp57 in hemostasis and thrombosis we generated a megakaryocyte/platelet specific knockout. Despite normal platelet counts and platelet glycoprotein expression, mice with ERp57-deficient platelets had prolonged tail-bleeding times and thrombus occlusion times, and defective activation of the αIIbβ3 integrin and platelet aggregation. The aggregation defect was corrected by addition of exogenous ERp57 implicating surface ERp57 in platelet aggregation. Platelet surface ERp57 protein and activity increased substantially with platelet activation. We conclude that platelet-derived ERp57 is required for hemostasis and thrombosis and platelet function.

Disclosures:

No relevant conflicts of interest to declare.
Effect of Increased Tissue Factor Pathway Inhibitor (TFPI) Levels On Factor Xa Inhibition and Global Hemostasis in the Presence of TFPI-Antagonistic Aptamer BAX 499.
Authors: Dockal Michael , Hartmann Rudolf , Knappe Sabine , Palige Michael , Kammlander Willibald , Kunckova Klara , Ehrlich Hartmut J. , Scheiflinger Friedrich
Published: October 2019

Abstract 2207

In a recent hemophilia clinical trial, patients' TFPI plasma levels substantially increased after intravenous or subcutaneous administration of TFPI-inhibitory aptamer BAX 499. To better understand the consequences of elevated plasma full-length (fl)-TFPI and the potency of BAX 499, the TFPI-inhibitory activity of a wide concentration range of BAX 499 was tested at levels up to 10 nM fl-TFPI in two model systems and in two global hemostatic coagulation assays.

Factor Xa (FXa) inhibition assay measured the influence of BAX 499 on the interaction between TFPI and FXa. BAX 499 at 1000 nM efficiently inhibited fl-TFPI within the physiological concentration (< 0.5 nM). However, with increasing TFPI, inhibition of TFPI by BAX499 became considerably less efficient, reflected by increasing EC50 and decreasing maximum inhibition. At substantially elevated TFPI concentrations (10 nM) EC50 increased ∼20-fold, with TFPI retaining ∼70% of its FXa inhibitory activity. Similar results were observed in a more complex reaction system (TF-FVIIa-catalyzed FX activation). These results suggest that BAX 499 is a partial inhibitor of TFPI, efficiently neutralizing TFPI at low concentrations and less efficiently inhibiting TFPI at high TFPI levels.

In thrombin generation assays with FVIII-inhibited plasma, a substantial excess of BAX 499 was required to compensate the anticoagulant activity of increasing fl-TFPI. Based on peak thrombin values, fl-TFPI levels that could be neutralized by a given BAX 499 concentration were determined. For example, a ∼50-fold excess of BAX 499 neutralized 0.2 nM TFPI added to FVIII-inhibited plasma, whereas a 140-fold excess was required for neutralization of 7.3 nM fl-TFPI added. An equivalent ROTEM experiment performed in FVIII-inhibited whole blood confirmed these findings.

In summary, the data showed that even a high concentration of BAX 499 (1 μM) was not able to counteract the strong inhibitory effect of 10 nM fl-TFPI on FXa, on TF-FVIIa-catalyzed FX activation, on thrombin generation in plasma and clot formation in whole blood, due to the partially inhibitory properties of BAX 499. The current study offers an explanation for increased bleeding tendency in hemophilia patients associated with the ≥25-fold elevated fl-TFPI plasma levels after BAX 499 administration. BAX 499 does not appear to be able to compensate the anticoagulant activity of increased TFPI levels caused by partial inhibition in patients treated with BAX 499.

Disclosures:

No relevant conflicts of interest to declare.
Functional Characterization of Baxter's Recombinant Human ADAMTS13 Drug Candidate.
Authors: Rottensteiner Hanspeter , Plaimauer Barbara , Schrenk Gerald , Schmidt Michaela , Gruber Bernadette , Schreiner Jutta , Turecek Peter L , Scheiflinger Friedrich
Published: October 2019

Abstract 2235

Baxter has developed a human recombinant (r) ADAMTS13 for treatment of patients with hereditary TTP. Here we characterize preclinical and clinical lots of rADAMTS13 with respect to their functional properties.

The ability of rADAMTS13 to degrade human full-length rVWF under moderate denaturing conditions was analyzed by two functional assays (VWF:CB and VWF:RCo activity) and by gel electrophoresis (multimer analysis and immunoblot analysis). In all assays, the extent of rVWF degradation served as a direct measure for ADAMTS13 activity.

VWF:CB activity of rVWF was reduced upon incubation with rADAMTS13, with only minimal differences between the individual batches. VWF:RCo activity of rVWF also declined after exposure to rADAMTS13, with no relevant differences between rADAMTS13 batches. Likewise, VWF multimer analysis demonstrated the disappearance of high molecular weight multimers upon incubation with rADAMTS13. Non-reducing SDS-PAGE followed by immunostaining using a polyclonal anti-human VWF antibody clearly showed the appearance of the C-terminal and N-terminal VWF fragments, with no visible differences in the intensity of these bands between preclinical and clinical Phase I rADAMTS13 batches.

Proteolytic activity of rADAMTS13 towards its multimeric VWF substrate was confirmed in two flow-based assays using full-length rVWF as substrate. One assay was based on the VenaFlux platform technology and determined the rADAMTS13 concentration-dependent decrease of the surface coverage of fluorescently-labeled platelets that had been perfused together with red blood cells and rVWF over an adhesive surface of a microcapillary. The other assay measured the extent of VWF cleavage product generation after subjecting a reaction mixture consisting of multimeric rVWF, lyophilized platelets and rADAMTS13 to fluid shear stress on a vortex mixer. Both assays revealed a high comparability between preclinical and clinical batches.

All assays thus demonstrated consistency between rADAMTS13 batches over different production stages. Importantly, comparability was also shown between the ADAMTS13 activity measured by the assays using full-length VWF as substrate and FRETS-VWF73 assay used for potency assignment of rADAMTS13.

Disclosures:

Rottensteiner: Baxter Innovations GmbH: Employment. Plaimauer:Baxter Innovations GmbH: Employment. Schrenk:Baxter Innovations GmbH: Employment. Schmidt:Baxter Innovations GmbH: Employment. Gruber:Baxter Innovations GmbH: Employment. Schreiner:Baxter Innovations GmbH: Employment. Turecek:Baxter Innovations GmbH: Employment. Scheiflinger:Baxter Innovations GmbH: Employment.
FDG-PET/CT Early After 90Y-Ibritumomab Tiuxetan Therapy Predicts Outcome in Relapsed or Refractory Indolent B-Cell Lymphoma
Authors: Okada Masaya , Fujimori Yoshihiro , Oku Naohiko , Tamekane Akira , Takafuta Toshiro , Nakajima Toshiyuki , Tokugawa Tazuko , Sawada Akihiro , Ishii Shin-ichi , Kaida Katsuji , Ikegame Kazuhiro , Soma Toshihiro , Ogawa Hiroyasu
Published: October 2019

Abstract 3648

Background:

Radioimmunotherapy (RIT) with 90Y-ibritumomab tiuxetan has been used for the treatment of relapsed or refractory indolent B-cell lymphoma. Early prediction of response to the therapy may offer the potential to identify patients who will benefit this therapy. We evaluated the efficacy of fluorine 18 fluorodeoxyglucose (FDG) combined positron emission tomographic-computed tomographic (FDG-PET/CT) imaging for assessment of therapy and predicting outcome in 90Y-ibritumomab tiuxetan radioimmunotherapy.

Methods:

Radioimmunotherapy with 90Y-ibritumomab tiuxetan was preformed in 52 patients with relapsed or refractory indolent B cell lymphoma (follicular lymphoma, 45; mucosa-associated lymphoid tissue lymphoma, 5; lymphoplasmacytic lymphoma, 2) at Hyogo College of Medicine Hospital from June 2009 through August 2012. FDG-PET/CT scanning was performed at two weeks and the later after 90Y-ibritumomab tiuxetan therapy.

Results:

In FDG-PET at 2 weeks after 90Y-ibritumomab therapy, 26 (50%) showed complete response (defined as early CR), 20 (38%) showed partial response (PR-2W) and 6 (12%) showed non response (NR-2W). Further follow-up revealed 10 later CR (8 CR out of 20 PR-2W and 2 CR out of 6 NR-2W), showing 69 %(36) of overall CR rate. Relapse was occurred in 4(17%) of 26 early CR and in 6 (60%) of 10 later CR, indicating significantly low relapse late in early CR (P= 0.0074, by Chi-square test). Patients with early CR (CR in 2 weeks after the RIT) showed significantly better progression free survival than those with later CR (P=0.0046, by Logrank test). In contrast, analysis at six weeks after the RIT showed 36 CR patients (10 of which eventually relapse), but failed to discriminate patients who had high risk of relapse.

Conclusion:

Our results suggest that CR assessed by FDG-PET/CT at 2 weeks after 90Y-ibritumomab tiuxetan therapy discriminates good responder to the RIT and predicts better progression free survival in relapsed or refractory indolent B cell lymphoma.

Disclosures:

No relevant conflicts of interest to declare.
Clinical Phase II Study of a Non-Anthracycline-Based Immunochemotherapy Regimen(R-GemOx) As First-Line Treatment in Elderly Patients with Diffuse Large B-Cell Lymphoma
Authors: Fan Lei , Wang Li , Xu Ji , Zhang Run , Wang Rong , Xu Wei , Li Jianyong
Published: October 2019

Abstract 3667

Introduction:

Anthracycline-based regimen(CHOP and CHOP-like regimens) is still the most widely used treatment of elderly patients with diffuse large B-cell lymphoma (DLBCL), but because of low rates of the overall response, high incidence of co-morbidities and treatment-related adverse effects, the prognosis of these patients remain dismal. Our aim was to develop a non-anthracycline-based immunochemotherapy regimen, rituximab combination with GemOx (gemcitabine and oxaliplatin) for elderly patients with DLBCL and investigate the safety and preliminary efficacy of this regimen.

Design and Methods:

This study was a prospective, single-arm clinical trial. Eligible patients was previously untreated, CD20 positive DLBCL patients diagnosed by biopsy, ≥70 years of age or ≥60 years with ECOG PS of ≥2. Therapeutic protocol of R-GemOx regimen was as follows: rituximab 375 mg/m2 i.v. on day 1; gemcitabine 1 g/m2 i.v. on day 2; oxaliplatin 100 mg/m2 i.v. on day 2, each cycle of treatment was administered every 14 to 21 days according to recovery of toxicities. Patients who responded to the treatment and did not have disease progression received up to six cycles of R-GemOx treatment.

Results:

15 patients were enrolled between July 2010 and April 2012 and 12 patients were eligible to evaluate. Patients were 41.7% (5/12) male with median age 73(range: 61–85 yrs), three patients (25.0%) were <age 70. ECOG PS ≥2 was 11/12 and the international prognostic score (IPI) enrolled to study was low risk group (0–1 score) 0/12, low intermediate group (2 score) 4/12, high intermediate (3 score) 2/12 and high risk group (4–5 score) 6/12. Evaluation of response was according to response criteria for lymphoma: The overall response rate (ORR) among treated pts was 75.0% (CR/CRu, 5; PR, 4); one patient (8.3%) had SD and 2 patients (16.7%) had PD. With a median follow-up of 7 (4–26) months, the median overall survival (OS) and the progression-free survival (PFS) have not been achieved. In terms of toxicities, most treatment related adverse events (AEs) were mild or moderate in severity. The most common grade ≥3 AEs were bone marrow depression (25.0%) and gastrointestinal complications (16.7%), no impairment of kidney and heart were observed.

Conclusions:

This is the first clinical study to investigate the safety and efficacy of R-GemOx regimen in elderly patients with DLBCL. Rituximab combination with GemOx appears highly active and favorable toxicity profiles in elderly patients with DLBCL requiring treatment, the encouraging preliminary data suggest R-GemOx is a promising first-line regimen for this subgroup of patients, and further evaluation is required.

Disclosures:

No relevant conflicts of interest to declare.
Bortezomib + DA-EPOCH-R Induction Therapy Followed by Maintenance Bortezomib Versus Observation in Newly Diagnosed Mantle Cell Lymphoma
Authors: Dunleavy Kieron , Grant Cliona , Hessler Julieanne , Miller Barry W. , Steinberg Seth M. , Pittaluga Stefania , Roschewski Mark , Jaffe Elaine S. , Wiestner Adrian , Wilson Wyndham H.
Published: October 2019

Abstract 3672

Background:

Bortezomib, a proteasome inhibitor, is effective in relapsed/refractory mantle cell lymphoma (MCL) and achieves responses in up to 50% of patients. While its mechanisms of action are broad, in MCL, brotezomib causes cell cycle arrest and induces apoptosis through oxidative/ER stress-mediated up-regulation of noxa. DA-EPOCH-R is an effective platform in MCL. We set out to assess the efficacy of bortezomib in combination with DA-EPOCH-R in newly diagnosed MCL and to assess the efficacy of maintenance bortezomib following induction.

Methods:

Treatment was divided into three parts: Part A - bortezomib (1.3 or 1.5 mg/m2) on days 1, 4, 8 and 11 for one cycle; Part B - bortezomib (1.3mg/m2) on days 1 and 4 of DA-EPOCH-R (as previously described) for 6 cycles; Part C patients in CR or PR after Part B and without significant neurotoxicity were randomized to maintenance bortezomib (1.3 mg/m2 d 1, 4, 8 and 11 q 8 wks for 18 mos) or observation.

Results:

Patient characteristics (n=43): median (range) age 58 (41–75); 45% were aged 60y or over; male sex 32 (73%); stage IV disease 100%. At 48 months (with 57 months median potential follow-up), PFS is 50% and OS is 80%. Twenty-four patients randomized to maintenance bortezomib (13) or observation (11) had a similar PFS (p=0.41) (see attached curve). Overall, 50% of patients had at least grade 2 neurotoxicity. At the recommended 1.3mg/m2 dose of bortezomib, 33% of patients required a dose reduction or discontinuation of drug. Other significant toxicities were neutropenic fever/infection in 8% of cycles and grade 3 or 4 thrombocytopenia in 34% of cycles.

Conclusions:

Bortezomib in combination with DA-EPOCH-R is an effective strategy in patients of all ages with MCL. Maintenance bortezomib (versus observation) does not improve the PFS. However, compared to our historical patients treated with DA-EPOCH-R alone, patients who received DA-EPOCH-R and bortezomib have a significantly better PFS at 48 months (50% versus 19%, respectively). These preliminary results suggest that bortezomib with immunochemothearpy may improve PFS. Accrual continues and responses will be correlated with translational end-points on biopsied tissue sampled before and after bortezomibalone (Part A).

Disclosures:

No relevant conflicts of interest to declare.