bluuebunny/crossref_metadata_2025 huggingface dataset:
Use of “high flow” Totally Implantable Central Venous Access Ports for Myeloablative Conditioning Followed by Allogeneic Stem Cell Transplantation: a Prospective Study.
Authors: Abdelkefi Abderrahman , Gaied Olfa Ben , Ladeb Saloua , Torjman Lamia , Lakhal Amel , Othman Tarek Ben
Published: October 2019

Abstract 4648

Objectives

A central venous access is always necessary for the management of patients receiving myeloablative conditioning followed by allogeneic stem cell transplantation (SCT). Tunnelled, cuffed silastic catheters are the device of choice for these patients. To our knowledge, there are no prospective studies investigating the use of totally implantable central venous access ports (TIAP) in patients receiving myeloablative conditioning followed by allogeneic SCT. The aim of this prospective study was to investigate the usefulness of a new “high flow” single lumen port device in these patients. <>Methods: Between December 2007 and July 2009, patients with haematological malignancies received a TIAP prior to allogeneic SCT. All patients received the same type of port [silicone, 10 French, high flow rate (3100ml/h), Reference 40010, Laboratoires Perouse, Ivry le Temple, France]. All devices were inserted under local anesthesia through direct puncture of the right subclavian vein using the Seldinger technique. A chest X-ray was always obtained to document correct positioning. All infusions, including the graft itself and all blood drawings, were performed via the port. Port-related bloodstream infection was defined according to Infectious Disease Society of America guidelines. All patients were examined by ultrasonography in case of clinical signs of thrombosis and systematically after discharge.

Results

Forty six TIAP were placed in 46 patients [median age: 25 years (20-41 years); 20 female and 26 male], and remained in place for a cumulative duration of 9756 days in situ (range 30-560 days). No pneumothorax occurred. Port-related bloodstream infection occurred in 2 cases (candida parapsilosis and staphylococcus aureus) [2/46, 4.3% ; 0.2 event per 1000 days]. These ports were removed. No port-related thrombosis occurred. No port-related deaths were observed.

In conclusion, the use of “high flow” totally implantable ports has resulted in a good option for long-term access to central veins and delivery of myeloablative conditioning followed by allogeneic SCT, in spite of severe neutropenia and increased risk of sepsis in this category of patients. Although multicentre randomized clinical trials are needed to define the optimal device in this clinical setting, the results of this prospective study support the wider use of TIAP in patients receiving myeloablative conditioning followed by allogeneic SCT.

Disclosures:

No relevant conflicts of interest to declare.
Population Pharmacokinetics Study of Tacrolimus in Chinese Hematopoietic Cell Transplant Patients.
Authors: Xue Ling , Rui Jian-Zhong , Zhang Yan , Miao Li-Yan
Published: October 2019

Abstract 4668

Background

The use of tacrolimus is complicated by its narrow therapeutic index and wide intra- and interpatient variability. The population pharmacokinetics (PPK) of tacrolimus have been reported in solid organ transplantation patients, but few investigation of tacrolimus PPK in hematopoietic cell transplant patients. Our study is to characterize the factors that influence tacrolimus population pharmacokinetics in Chinese hematopoietic cell transplant patients.

Methods

Sixty-eight patients undergoing hematopoietic cell transplant were enrolled. The database (671 data points) collected from the patients was analyzed by using the nonlinear mixed-effect model (NONMEM) program. Patients were administered oral or/and intravenous tacrolimus as part of immunosuppressive regimen that also included corticosteroids. And the robustness and accuracy of the model were assessed using bootstrap. A number of covariates including demographic characteristic, biochemical and hematological index, drug combination, IOV (inter-occasion variability) and other factors (e.g. Cause, POD, the type of transplantation and the sources of donor) were evaluated for their influence on the tacrolimus population pharmacokinetic parameters.

Results

The basic model was selected a one-compartment pharmacokinetic model with first-order absorption and elimination. The typical values of tacrolimus CL, V, F were 12.1 L·h-1, 686 L, 42.2%, respectively. And the inter-individual variations of CL, V, F were 23.5%, 96.4%, 43.8%, respectively. The absorption rate constant was fixed 4.3 h-1. The residual error between observed and model-predicted concentration was 3.03 ng·ml-1. The covariates that influenced the population pharmacokinetics parameters of tacrolimus included inhibitor (INHI), post operation days (POD), age and hemoglobulin (HGB) in the final regression model. The IOV of CL, V, F were 22.2%, 6.23%, 30.3%, respectively.

Conclusion

The covariates included INHI, POD, AGE, HGB and IOV would significantly influence tacrolimus population pharmacokinetics parameters in hematopoietic cell transplant patients.

Disclosures:

No relevant conflicts of interest to declare.
High Id1 Expression Is Associated with Poor Prognosis in 237 Patients with Acute Myeloid Leukemia.
Authors: Hirsch Pierre , Tang R. , Lapusan Simona , Marzac Christophe , Marie Jean-Pierre , Legrand Ollivier
Published: October 2019

Abstract 4687

Inhibitors of differentiation (Id) or DNA binding helix-loop-helix (HLH) proteins are a group of dominant inhibitors of basic HLH transcriptional factors, which act as positive regulators of cell growth or promote excessive proliferation, and also protect cells against drug-induced apoptosis in mammalians. Recently, Id1 has been identified as a common downstream target of several constitutively activated oncogenic tyrosine kinase(TK), such as FLT3/ITD, in leukemia cells. We analyzed Id1 expression as possible prognostic factor in 237 AML patients. High Id1 expression was associated with older age(p=0.009); and with Ftl3/ITD(p=0.003). However, 61% of the patients in the group of FLT3- AML were Id1+, suggesting that other TK are involved. In whole population, high Id1 expression independently predicted shorter disease free survival (DFS)(p=0.05), and overall survival (OS)(p=0.003). In the subgroup of young patients (age'60 years) with normal cytogenetic, Id1+ was, in multivariate analysis, associated with lower CR rates(p=0.02), shorter DFS(p=0.02) and OS(p=0.006).

In conclusion our data provides a new molecular marker for refining the risk classification of AML especially in young patients with normal cytogenetic. Id1- patients with normal cytogenetic should be classified as favourable-risk leukemia. Id1, as a downstream target of constitutively activated TK, could be a suitable candidate for targeted therapy.

Disclosures:

No relevant conflicts of interest to declare.
Targeting Tissue Factor to Tumor Vessels. Experimental Results and First-in-Man Experience.
Authors: Berdel Wolfgang E , Kessler Torsten , Schwöppe Christian , Liersch Ruediger , Persigehl Torsten , Bremer Christoph , Schliemann Christoph , Mesters Rolf
Published: October 2019

Abstract 469

Activation of blood coagulation in tumor vessels with subsequent tumor infarction is an experimental strategy in cancer therapy. We have fused different targeting peptides, including GRGDSP (RGD), GNGRAHA (NGR) and 5 cyclic derivates to the C-terminus of truncated tissue factor (tTF) to preferentially target tumor vessel endothelial cell integrins such as αvβ3 or aminopeptidase (CD13). tTF fusion proteins were expressed in E. coli, purified and refolded. Molecular integrity of the fusion proteins was evaluated by SDS-PAGE, immunoblotting and mass spectrometry. Subsequently, the tTF-fusion proteins were tested for biological activity with the following results: They retained the thrombogenic activity of tTF as measured by factor × activation in vitro. When tested with their respective target molecules either in a purified preparation or present on growing endothelial cells, there was specific binding. In vivo studies with human tumor xenograft models in nude mice showed either significant inhibition of tumor growth or regression of established tumors of different histologies (e.g. lung, breast, melanoma, sarcoma) by systemic application of the tTF-fusion proteins in contrast to controls including non-targeted tTF. Histology of the tumors treated with tTF fusion proteins revealed thrombotic occlusion of vasculature and blood pooling. Contrast-enhanced magnetic resonance imaging (MRI) of the tumors in vivo before and shortly after application of tTF-RGD and tTF-NGR showed a significant reduction of tumor perfusion. Degree of reduction correlated with in vivo tumor response. Toxicity studies showed acceptable therapeutic range and at therapeutic dosage there was no thrombo-embolic event in histology of normal organs, such as lung, heart, liver, and kidney. After upscaling production to amounts sufficient for clinical use, we have treated the first cancer patients with tTF-NGR. MRI studies even at the lowest dose (1 mg/kg i.v.) given showed reduction of tumor perfusion with no side effects. Targeted infarction of tumor vasculature with tTF fusion proteins may be promising as cancer therapy and should be further studied.

Disclosures:

Berdel: private: patent application on targeting tissue factor. Mesters:private: patent application on targeting tissue factor.
SET-NUP214 Fusion Gene at Chromosome 9q34 Contributed to the Development of Not Only T-ALL, but Also Primary and Secondary AML.
Authors: Shin Myung-Geun , Kim Da-Woon , Kim Hye-Ran , Cho Duck , Kee Seung-Jung , Kim Soo-Hyun , Shin Jong-Hee , Suh Soon-Pal , Ryang Dong-Wook
Published: October 2019

Abstract 4696

SET-NUP214 rearrangement at chromosome 9q34 is an extremely rare recurrent cryptic deletion resulting from del(9)(q34.11q34.13) mainly found in T-cell acute lymphoblastic leukemia (T-ALL). A multiplex RT-PCR system (HemaVision; DNA Technology) covering 28 leukemic fusion transcripts was applied to 271 bone marrow (BM) samples obtained from acute leukemia patients at initial diagnosis to investigate new pathophysiologic role of SET-NUP214 fusion gene. Out of 271 samples, we identified three cases (1.1%) harboring SET-NUP214 rearrangement – each case of adult T-ALL, pediatric acute myeloid leukemia (AML) from chronic myeloid leukemia (CML) and adult de novo AML with minimal differentiation (M0). Adult T-ALL occurred in 38-year-old man presented with dizziness and hepatomegaly. Conventional cytogenetic study disclosed normal karyotype and existence of SET-NUP214 fusion gene generated by del(9)(q34.11q34.13) was confirmed by RT-PCR, FISH and direct sequencing. Hematological remission was achieved after treatment of induction chemotherapy but during 7 months follow-up, SET-NUP214 fusion gene was steadily detected by RT-PCR. 13-year-old female with AML M0 converted from CML after treatment of imatinib for 2 months was the second case. Leukemic cells showed negative myeloperoxidase (MPO) and sudan-black B (SBB) staining, positive reaction against CD13, CD33, CD117 and MPO monoclonal antibodies by flowcytometry-based immunophenotyping. Major BCR-ABL1 rearrangement and SET-NUP214 fusion genes were simultaneously detected by RT-PCR, FISH and direct sequencing. She died at 38 days after diagnosis during the chemotherapy. The fusion protein contributed to the leukemogenesis by promoting expression of HOXA cluster genes. Last case was a de novo AML M0 in 39-year-old male. Cytochemical stains of leukemic cells on MPO and SBB were negative and immunophenotyping studies were positive for CD34, HLA-DR, CD13 and CD33. SET-NUP214 rearrangement was detected by multiplex RT-PCR. After treatment of induction and consolidation chemotherapy, he received allogeneous sibling stem cell transplantation (SCT), then complete chimerism was achieved. In clonclusion, SET-NUP214 rearrangement is associated with the development of not only T-ALL, but also primary and secondary AML via aberrant expression of HOXA cluster genes.

Disclosures:

No relevant conflicts of interest to declare.
Interphase Fluoroscence in Situ Hybridization Improves the Detection Rate of CNS Involvement in Childhood Acute Lymphoblastic Leukemia.
Authors: Park Seungman , Lee Sangho , Kang Hyoung Jin , Shin Hee Young , Ahn Hyo Seop , Kim Hyun Kyung , Lee Dong Soon , Cho Han Ik
Published: October 2019

Abstract 4714

Background

Morphologic examination of CSF cytospin is the standard method in the detection of CNS involvement of childhood acute lymphoblastic leukemia (ALL). However, the result widely varies depending on the status of cell degeneration caused by cytospin procedure. We performed interphase fluoroscence in situ hybridization (FISH) on CSF cytospin cells to examine its potential usefulness in detecting malignant cell population in the CSF.

Materials and Methods

The study used 19 CSF cytospin specimens of the childhood ALL patients under treatment. All of them had cytogenetic abnormalities in the bone marrow nuclear cells at the initial diagnosis. None of the patients showed malignant cells in morphologic examination of the cytospin. Interphase FISH was performed to detect recurrent genetic abnormalities including rearrangement of BCR/ABL, TEL/AML1, MLL and IgH rearrangement, and p16 deletion using LSI BCR/ABL dual color, dual fusion translocation probe, LSI TEL/AML1 ES dual color translocation probe, LSI MLL dual color, break apart rearrangement probe, LSI IGH dual color, break apart rearrangement probe, and LSI p16/CEP 9 dual-color probe (Abbot Molecular Diagnostics). FISH signals were interpreted using automated slide scanning system (Metafer 4 system) and manually read for confirmation.

Result

Two patients had the same cytogenetic abnormalities in CSF cytospin as observed in the bone marrow mononuclear cells: one patient had 4 copies of AML1 gene, suggesting tetrasomy 21 or multiple fusion partner gene other than chromosome 12, both in bone marrow and in CSF; the other had 3 copies of p16 gene including one with low signal intensity both in bone marrow and in CSF. Ten patients did not have any cytogenetic abnormalities. We could not interpret the result in the other 7 patients due to low cell count and/or low signal intensity.

Conclusion

Out of 19 who were diagnosed not to have malignant cell in CSF in morphologic examination, 2 patients had cells with the same clonal abnormalities in the CSF as bone marrow. Our result indicates that FISH study would be helpful in evaluating CSF involvement of hematologic malignancies including ALL. In order to improve the performance, it is required to prevent degeneration of the cells by prompt specimen delivery and slide preparation.

Disclosures:

No relevant conflicts of interest to declare.
The Use of ABL and GUS as Control Genes for Normalizing Quantification of Fusion Transcripts by Real Time PCR: Our Experience of More Than 1000 Samples.
Authors: Lassoued Mohamed Amin Ben , Beaufils Nathalie , Loundou Anderson Dieudonné , Arnaud Christiane , Gabert Jean
Published: October 2019

Abstract 4727

Introduction

Real-time quantitative PCR (RT-PCR) is a sensitive and accurate tool for monitoring leukemic patients by amplification of fusion gene (FG) transcripts.

In order to correct variations in RNA quality and quantity and to calculate the sensitivity of each measurement, a control gene (CG) transcript should be amplified in parallel to the FG transcript.

In our laboratory two CG are used: Abelson (ABL) and beta-glucuronidase (GUS).

Clinical application for monitoring the treatment of chronic myelogenous leukemia (CML) patients with tyrosine kinase inhibitor makes the quantification of which CG to use very important into the clinical practice today.

Objective

The aim of this study was to assess the degree of agreement between the GC, compare the expression of GC (type and time of sampling and pathology), try to redefine the limits between the 4 groups of quality (very good, good, poor and very poor quality samples) and to focus on CML patients.

Materials and methods

A total of 1087 samples (peripheral blood and bone marrow) were obtained from 475 patients from January 2005 to December 2007. This study includes patients with acute myeloid leukaemia (AML), acute lymphoblastic leukemia (ALL), chronic myeloid leukaemia (CML) and Hypereosinophilic syndrome (HES).

We collected 366 samples of 41 CML patients between 2002 and March 2009 including 32 CML at diagnosis.

RT-PCR analysis was performed using the EAC protocols.

Results

Comparison between CML, AML, ALL and HES patients showed that ABL and GUS expression did not differ significantly (Bland and Altman method).

ABL and GUS gene expressions did not differ significantly between CML patients at diagnosis and the others.

Conclusion

Based on the literature and our local experience of more than 1000 samples, we propose to use only ABL as CG in routine practice in onco-hematology particularly for CML patients at diagnosis.

Disclosures:

No relevant conflicts of interest to declare.
High Dose Cyclophosphamide for Moderate to Severe Refractory Multiple Sclerosis: 2-Year Follow-up (investigational new drug No. 65863).
Authors: Gladstone Douglas
Published: October 2019

Abstract 4744

Background

High dose cyclophosphamide (HDC) is a chemotherapy treatment designed to eradicate autoreative B- and T-cells responsible for lymphocyte-mediated autoimmune illness, while sparing the pluripotent blood stem cell of any ill effects. Multiple sclerosis (MS) is the most common inflammatory and demyelinating immune-mediated disorder of the central nervous system in young adults.

Methods

Patients with moderate to severe, refractory MS, defined as an Expanded Disability Status Scale (EDSS) score of 3.5 or higher after 2 or more Food and Drug Administration-approved disease-modifying agents, received 200 mg/kg of cyclophosphamide over 4 days. For the following 2 years, quarterly EDSS score evaluations and biannual brain magnetic resonance imaging and neuro-ophthalmologic evaluations were obtained.

Results

15 patients were evaluated for clinical response. During follow-up, no patients increased their baseline EDSS score by more than 1.0. EDSS score stability or decrease was realized in 5 of 7 (71%) patients with relapsing remitting MS and 5 of 8 (62%) patients with secondary progressive MS patients. 4 patients required additional immunomodulatory treatment after treatment. Neurologic improvement involved changes in gait, bladder control, and visual function. Treatment response was seen regardless of the baseline presence or absence of contrast lesion activity.

Conclusions

HDC can effectively decrease symptoms, stop disease progression, and allow for disability regression in RR and SPMS patients. The most appropriate candidates for HDC, its duration of benefit and the potential need for prophylactic preventative immune manipulation after HDC all require further investigation.

Disclosures:

Off Label Use: cyclophosphamide: IND# 65863.
Elevated Heat Shock Protein 70 (HSP70) Serum Levels After Tumor Radiofrequency Ablation Might Represent a Favorable Prognostic Marker.
Authors: Haen Sebastian P. , Pereira Philippe L. , Clasen Stephan , Schmidt Diethard , Boss Andreas , von Herbay Alexandra , Kanz Lothar , Rammensee Hans-Georg , Gouttefangeas Cecile
Published: October 2019

Abstract 4748

Introduction

Radiofrequency (RF) ablation is a safe and effective method to locally treat malignancies of the liver, kidney and more recently lung. For the treatment of small tumors, long term results are comparable with outcome after surgical resection and better than current chemotherapies. RF-ablation has now been reported to induce heat shock protein (HSP) expression on sublethally damaged hepatocytes. HSPs are potent alarm signals for the immune system causing i.e. activation of T cells and dendritic cells. In addition to triggering immune responses directed against HSPs themselves, HSPs can carry antigens derived from those cells of which HSPs originated and thereby elicit specific immune reactions against these cellular epitopes.

Patients and Methods

We here analyzed serum samples of 22 patients to evaluate the systemic release of HSP70 using a specific HSP70 ELISA. Patients were followed for a median of 30 months until death or three years after the last RF-treatment. As a control group, sera from 20 patients undergoing diagnostic liver biopsy were screened.

Results

The median increase of HSP70 in serum comparing levels before and one day after the procedures was 1.6-fold in the RF-ablation group, while no soluble HSP-70 could be detected in the control group before and after the biopsy. Nine patients showed a higher than two-fold increase of HSP70 in the serum after ten sessions of RF-ablation compared to levels prior to treatment. No correlation of HSP70 levels with tumor volume, the ablated non-malignant tissue or the total size of the necrosis was observed. In the subgroup of patients with elevated HSP-70 serum levels, 67% (6 out of 9) were alive at the end of follow up with only one suffering from progressive disease. In contrast, 61% (8 out of 13) of the other patients suffered from progressive disease or had deceased at the end of follow-up.

Conclusion

RF-ablation not only induces cellular HSP expression but also can lead to a systemic release of serum HSP70. Patients with strong increases in HSP-70 serum levels tended to have a better survival than patients without induction of HSP-70 in the serum. Thus, immunomodulation might represent an additional beneficial effect of RF-ablation besides local tumor destruction, since HSPs are strong stimuli for the immune system and can mediate tumor-specific T cell immunity.

Disclosures:

No relevant conflicts of interest to declare.
First Use of Temozolomide in Primary Intra-Ocular Lymphoma (PIOL): a Very Well Tolerated and Potentially Effective Treatment.
Authors: Mesmoudi Mohamed , Cassoux Nathalie , Leblond Veronique , Merle-Beral Helene , Baudet Sylvie , Choquet Sylvain
Published: October 2019

Abstract 4760

Background

PIOL is a rare subset of primary central nervous system lymphoma (PCNL), representing 5 to 20% of PCNL. It is usually an aggressive diffuse large B-cell lymphoma. There is no consensus on treatment procedures, classical attitudes are systemic chemotherapy (SC), radiotherapy or intraocular injection of methotrexate (IM). Relapse rate is high, with more than 10% of cases relapsing in brain. New treatments are necessary, especially with a good tolerance profile. Temozolomide (Te) has some efficiency on PCNL and seems to be a potential candidate.

Methods

we retrospectively analyzed PIOL treated by Te in our unit. Inclusion criteria were a diagnosis established on cytological and molecular analysis after vitrectomy or anterior chamber puncture, and the absence of brain or meningeal localization. Interleukin 10 (IL10) and 6 (IL6) dosages were made by enzyme-linked immunosorbent assay on ocular material. IL10/6 ratio >1 is a specific and sensitive test for PIOL diagnosis and IL10 kinetic a good marker of response in parallel with clinical examination. Treatment consisted in Te at 150mg/m2 orally 5 days per month, in monotherapy, without corticosteroid use.

Results

four patients were analyzed, 3 males and one female. All received systemic chemotherapy with at least high dose methotrexate in first line, two with addition of high dose cytarabin. First patient, aged 84 at diagnosis, relapsed 15 month after CR and received Te for three months until here death, probably by CNS lymphoma. Second patient, aged 65, progressed under SC and subsequently received IM without any success; Te was used in third line but no effect was seen. Third patient, aged 48, had a partial response after first line treatment and received SC with high dose cytarabin and stem cell transplantation; after CR he relapsed 33 months later and then received Te, partial response was clinically confirmed at 3 months and is stable at 6 months. Last patient, age 71, had a localized PIOL 3 years after the treatment of a follicular lymphoma; PIOL relapsed 6 month after the first line and Te began; clinical response was spectacular, with a vision normalization in less than one month, a clinical CR confirmed at two months, IL10 dosages were at more than 1500 pg/ml in both eyes before Te, at 22 pg/ml (left eye) after one month and 202 pg/ml (right eye) after two months, response is stable at 5 months. No toxicity appeared during treatment except for hematological grade I or II.

Conclusion

Temodal is a safe and effective treatment of PIOL after classical chemotherapy. Longer follow-up and larger studies are necessary to confirm these data

Disclosures:

No relevant conflicts of interest to declare.
Gemcitabine Is An Active Agent in the Treatment of T Cell Non Hodgkin Lymphomas.
Authors: Jajeh Ahmad
Published: October 2019

Abstract 4771

Approximately 12- 15% of non- Hodgkin lynphoma NHL are caused by malignant Tcell lymphocytes. The success of the treatment in the aggressive form has lagged behind that of B -Cell in term of poor resonse and durability. Peripheral T-Cell lymphoma PTCL and cutaneous T-Cell lymphoma CTCL are two major charachterized classifiction in the World Health Organization based on their morphology, growth patern and genetics. Stem cell transplant and high dose chemotherapy have been associated with long term response rate of 45%. However this treatment is not well tolerated and not feasible for many patients. Other theraputic options include cytotoxic drugs CHOP,CVP regimen, purine analogues, Denileukin diftitox, Histone deacetylation inhibitors and novel antifolates drugs. In this abstract we will show our experience with gemcitabin an active antimetabolite as a single agent or in combination with other active drugs. Eight patients with PTCL, five with visceral stage of mycosis fundoides( one patient with HIV infection), two with refractory anaplastic Ki positive NHL and one with angioimmunoblastic type. All patients failed a minimum two lines of therapy. Mean age 58 years( range 28-75). Eight blacks, two whites, four hispanics and one asian. Meduim cycles given are four. The dose range 800-1000 mg/M2, given weekly x3 every 28 days cycle. Overall response rate is 85%. Complete response rate CR in eight patients ( three PTCL, three mycosis fungoides and two anaplastic large cell NHL). Partial response PR in four and stable disease in one. Median duration of response is nine months, range six to two years. Median time for response is six weeks. In conclusion:Gemcitabine is an active drug in T-Cell lymphomas particularly when used in combination with other active agents. Maintenance dosing or retreatment with this drug should be investigated.

Disclosures:

No relevant conflicts of interest to declare.
Identification of a Predictive Factor for Reversible Neurological Adverse Events in a Subset of Non-Hodgkin Lymphoma Patients Treated with CD19-Specific BiTE ® Antibody Blinatumomab.
Authors: Zugmaier Gerhard , Nagorsen Dirk , Klinger Matthias , Bargou Ralf C , Goebeler Mariele , Viardot Andreas , Baeuerle Patrick A , Kufer Peter
Published: October 2019

Abstract 4793

Blinatumomab is a CD19/CD3-bispecific antibody construct of the bispecific T cell engager (BiTE®) class showing as single agent a high rate and duration of responses in patients with relapsed non-Hodgkin lymphoma (NHL) and B-precursor acute lymphocytic leukemia (ALL). Blinatumomab has a favorable safety profile with exception of a subset of patients developing neurological adverse events (AEs) during the first days of treatment, such as confusion, speech impairment or cerebellar symptoms. Thus far, all relevant neurological AEs (11 out of 48 patients) were transient, fully reversible and resolved without sequelae within 3 to 72 hours after stop of infusion. In no case, pathological findings were seen upon cranial magnetic resonance imaging. Despite treatment discontinuation, 4 patients with neurological AEs have achieved an objective lymphoma remission. Analysis of cerebrospinal fluid (CSF) taken within hours after stop of infusion showed detectable levels of blinatumomab in the majority of affected patients, while in one patient without neurological symptoms no blinatumomab was detectable in CSF during infusion. Moreover, increased levels of albumin and T lymphocytes in CSF support a disturbance of the blood brain barrier (BBB) as a possible underlying event. Analyses of patient serum samples for angiopoetin-2 and S100b are ongoing to investigate whether levels of the endothelial stress and BBB integrity marker, respectively, correlate with neurological AEs. In a retrospective analysis of 39 NHL patients, a baseline B cell to T cell (B:T) ratio in peripheral blood at or below 1:10 was identified as the only predictive factor for the subsequent occurrence of neurological AEs. The predictive value was then prospectively confirmed in 8 additional patients. Of note, ALL patients –despite very low B:T ratios– rarely showed neurological AEs, which may relate to previous intrathecal chemotherapy depleting target cells in the brain. Potential mechanisms for the neuroprotective effect of peripheral B cells are being investigated. In conclusion, we identified a simple measure to prospectively identify patients at risk of developing neurological AEs after onset of blinatumomab treatment. Mitigating measures are currently tested in these high-risk patients in order to avoid discontinuation of treatment.

Disclosures:

Zugmaier: Micromet: Employment, Equity Ownership. Nagorsen:Micromet: Employment, Equity Ownership. Klinger:Micromet: Employment, Equity Ownership. Bargou:Micromet: Consultancy, Patents & Royalties. Baeuerle:Micromet: Employment, Equity Ownership. Kufer:Micromet: Employment, Equity Ownership, Patents & Royalties.
Effect of Protein Kinase C β Specific Inhibition On Acute Lymphoblastic Leukemia Cell Lines.
Authors: Saba Nakhle S , Safah Hana F , Levy Laura S
Published: October 2019

Abstract 4817

Acute lymphoblastic leukemia (ALL) is the most common leukemia in children and accounts for 20% of acute leukemia in adults. The intensive induction–consolidation–maintenance therapeutic regimens used currently have improved the 5-year disease free survival to around 80% in children and to 25%-40% in adults. The poorer response in adults is basically due to the inability to tolerate the intensive chemotherapy, and to the biology of adult disease which is associated with poor-risk prognostic factors. In the present era of target-specific therapy, PKCβ targeting arose as a new, promising, and well tolerated treatment strategy in a variety of neoplasms, especially in B-cell malignancies. It showed encouraging results in preclinical and clinical studies involving chronic lymphocytic leukemia, diffuse large B-cell lymphoma and multiple myeloma. PKCβ plays a major role in B-cell receptor signaling, but studies describing the role of PKCβ in B-cell ALL are lacking. In the present study, we measured the sensitivity of a variety of B-cell ALL cell lines to PKCβ specific inhibition. Three cell lines were studied: RS4;11 (characterized by the t(4;11) chromosomal abnormality), TOM-1 (characterized by the t(9;22) chromosomal abnormality), and REH (characterized by the t(12;21) chromosomal abnormality). Cells were tested for PKCβ1 and PKCβ2 expression by immunoblot. Cell viability was measured when PKCβ-specific inhibitor at concentrations of 1, 2.5, 5, 10, 20 and 30 μM was added for 48 hours in the presence of 10% fetal bovine serum (FBS). MTS assay was performed to quantify cell viability. Results showed that all three cell lines express PKCβ1 and PKCβ2. Treatment with PKCβ-specific inhibitor resulted in a dose-dependent inhibition of cell proliferation; Sensitivity was evident at 1 μM for RS4;11 cell line, and at 2.5 μM for TOM-1 and REH cell lines, with 10% cell growth inhibition; Growth inhibition increased to 90% for all cell lines at an inhibitor concentration of 30 μM. These results indicate that PKCβ plays an important role in the malignant process in B-cell ALL, and suggest that PKCβ targeting should be considered as a potential treatment, whether in combination with the current regimens used or as a single agent monotherapy. Ongoing studies in our lab will detail the mechanism of PKCβ and adverse cytogenetics like t(4;11) and t(9;22).

Disclosures:

No relevant conflicts of interest to declare.
AICAR Induces Apoptosis in Chronic Lymphocytic Leukemia Cells through the Mitochondrial Pathway and Independently of p53 Pathway and AMPK Activation.
Authors: Gil Joan , Santidrián Antonio F , González-Gironès Diana M , Iglesias-Serret Daniel , Coll-Mulet Llorenç , Cosialls Ana M , de Frias Mercè , Campàs Clara , González-Barca Eva , Alonso Esther , de Sevilla Alberto Fernandez , Viollet Benoit , Pons Gabriel
Published: October 2019

Abstract 4819

AICAR (5-aminoimidazole-4-carboxamide riboside or acadesine) induces apoptosis in different cell types including chronic lymphocytic leukemia (CLL) cells. Here, we have analyzed the mechanisms involved in AICAR-induced apoptosis in CLL cells. AICAR induced apoptosis through the mitochondrial pathway, since inhibition of caspase-8 did not protect CLL cells from AICAR-induced apoptosis and caspase inhibition did not alter cytochrome c release induced by AICAR. AICAR induced a significant increase in the mRNA levels of the proapoptotic BH3-only genes BIM, BNIP3, BNIP3L, HRK, MOAP1, and NOXA. These changes were AICA ribotide (ZMP) accumulation-dependent and adenosine monophosphate-activated protein kinase (AMPK) activation-independent in CLL cells. Furthermore, AICAR induced the accumulation of NOXA protein in all CLL samples and BIM protein in about half of these samples, without modifying the levels of other BCL-2 family proteins analyzed. Importantly, AICAR induced apoptosis irrespective of the tumor suppressor TP53 and ataxia telangiectasia mutated status in CLL cells. AMPK activation with phenformin or A-769662 did not induce apoptosis in CLL cells. Finally, AICAR induced apoptosis in B lymphocytes from AMPKa1−/− mice. Taken together, our results demonstrate that AICAR induces apoptosis in B lymphocytes through the mitochondrial pathway by an AMPK- and p53-independent mechanism.

Disclosures:

Gil: Advancell-In Vitro Cell Technologies S.L.: Patents & Royalties, Research Funding. Campàs:Advancell-In Vitro Cell Technologies : Employment, Patents & Royalties.
The Study of Bortezomib Alone or in Combination with Arsenic Trioxide on Multidrug Resistance of HL60/ADR cells.
Authors: Meng Fanyi , Huang Ming
Published: October 2019

Abstract 4823

INTRODUCTION

Proteĉ¢msome inhibitor bortezomib has used in treatment of hematology malignancies widely. We found it may reduce cell survival rate of HL60/ADR cell line and induce cell apoptosis in previous study. Now we are to investigate the effect of bortezomib alone or combined with arsenic trioxide (As2O3) on reversing multidrug resistance of HL60/ADR cell line and the possible machines.

METHODS

HL60/ADR cells were incubated with bortezomib at different doses alone and in combination with As2O3. The proliferation ratio was observed by MTT assay. Cell apoptosis was studied by fluorescence microscopy and flow cytometry. Intracellular concentration of daunorubicin (DNR) and multidrug resistance related protein-1 (MRP1) was determined by flow cytometry. P65Ap-p65Abcl-2AbaxAcaspase-3Acaspase-9APARP proteins were determined by western blot.

RESULTS

In bortezomib-treated tumor cells, inhibition rate increased in time- and dose-dependently, as well as apoptotic cells. Compared to bortezomib alone, combination with As2O3 inhibited the proliferation and induced the apoptosis of HL60/ADR cells more evident. Bortezomib can enhance the intraceflular accumulation of DNR and decrease MRP1 in HL60/ADM cells. The dual combination of As2O3 with bortezomib presents a superior anticancer and MRP1-decreased efficiency to either one of the drugs alone. Bortezomib can also elevate the expression of bax, caspase-3, caspase-9, PARP, and decelerate the expression of bcl-2, NF-κB p65, p-p65.

CONCLUSIONS

Bortezomib can reverse multidrug resistance of HL60/ADR cells and decrease the expression of MRP1 in cells. When combined with As2O3, it appears to synergistic effects. Its mechanisms might be associated with the inhibition of NF-κB activation, also with inhibiting anti-apoptosis proteins, boosting pro-apoptosis proteins, with followed activating caspase pathway.

Disclosures:

No relevant conflicts of interest to declare.
Re-Exposure of Cord Blood (CB) to Non-Inherited Maternal HLA Antigens (NIMA) Improves Transplant Outcome in Patients (pts) with Hematologic Malignancies and May Reduce Relapse.
Authors: van Rood Jon J , Stevens Cladd E , Smits Jacqueline , Carrier Carmelita , Carpenter Carol , Scaradavou Andromachi
Published: October 2019

Abstract 661

CB hematopoietic stem cell transplantation (CBT) can be successful even if donor and recipient are not fully matched for human leukocyte antigens (HLA). This may result, at least in part, from tolerance-inducing events during pregnancy, but this concept has not been tested to date. Hence we analyzed the impact of fetal exposure to NIMA of the HLA-A, -B antigens or -DRB1 alleles on the outcome of 1121 pts with hematologic malignancies. All pts received single CB units provided by the NYBC, for treatment of ALL (N=451), AML (N=376), CML (N=116), MDS (N=79), other (N=99); 22% were transplanted in advanced stage. Median age was 9.7 years (range: 0.1-67); 29% of recipients were >16 years. Most pts (96%) received myeloablative cytoreduction. Sixty-two pts received fully matched grafts while 1059 received units mismatched (MM) for one or two HLA antigens. Of these, 79 (7%) had a MM antigen which was identical to a donor NIMA (Example: Pt: A1, A3; CBU: A1, A2; mother-CBU: A1, A3; A3 is NIMA). NIMA match was found in 25 recipients with one HLA MM and 54 of those with two MM. The NIMA match was identified after the transplant and was not used in unit selection. In multivariate analyses, NIMA matched transplants (NMTs), showed faster neutrophil recovery (RR=1.3, p=0.043), even for grafts with cell dose <3×107 (RR=1.6, p=0.053). There was no difference in the incidence of acute (grade II-IV) or chronic GvHD. 3-year relapse risk (cumulative incidence 22%) was reduced compared to 1 or 2 HLA MM no NIMA matched transplants, especially in pts with myelogenous malignancies given units with 1 HLA MM (RR=0.2, p=0.074). Further, 3-year transplant-related mortality was reduced (RR=0.7, p=0.034), particularly in pts ≥5 years old (RR=0.5, p=0.006), as was the 3-year overall mortality (RR= 0.7, p=0.029 and RR=0.6, p=0.015, respectively). As a result, in the NMTs, treatment failure (relapse or death) was significantly lower, particularly in pts ≥5 years (RR=0.7, p=0.019) and DFS was significantly improved (figure) and was similar to that of the 0 HLA MM group. These findings are the first indication that donor exposure to NIMA can improve post-transplant survival in unrelated CBT and might reduce relapse. We propose to include the NIMA of CB units in search algorithms. Thus, for pts lacking fully HLA matched grafts, HLA MM but NIMA matched CB units could be selected preferentially, since no adverse effects were seen. This strategy of selecting HLA MM grafts with optimal outcome effectively “expands” the current CB Inventory several-fold.Patient GroupNRR(95% Cl)p value0 MM360.5(0.3–0.8)0.0051 MM / NIMA Match180.4(0.2–0.9)0.0262 MM / NIMA Match400.8(0.5–1.2)0.3091 MM / No NIMA Match229reference group2 MM / No NIMA Match4871.1(0.9–1.3)0.365

Disclosures:

No relevant conflicts of interest to declare.
Identification of New Recurrent Multiple Small Interstitial Deletions Affecting Genes Coding for Kinases in Chronic Lymphocytic Leukemia (CLL): a New Pathogenic Mechanism?.
Authors: Rinaldi Andrea , Mian Michael , Rossi Davide , Forconi Francesco , Deambrogi Clara , Spina Valeria , Cencini Emanuele , Sozzi Elisa , Drandi Daniela , Ladetto Marco , Santachiara Rita , Marasca Roberto , Chigrinova Ekaterina , Cavalli Franco , Gattei Valter , Zucca Emanuele , Gaidano Gianluca , Kwee Ivo , Bertoni Francesco
Published: October 2019

Abstract 672

BACKGROUND:

CLL is the most common adult-onset leukemia in the Western world. The most common known genetic lesion is the 13q14.3 deletion targeting MIR15/MIR16. We applied a very high resolution array to identify new genetic lesions in CLL.

METHODS:

266 CLL samples were analyzed with Affymetrix Human Mapping 6.0 arrays, comprising over 1,8 million probes with a median distance of less than 1 Kb. Copy number was inferred using the circulary binary segmentation (CBS) algorithm. Minimal common regions (MCR) were defined using a modified version of the algorithm by Lenz et al. (PNAS 2008), specifically altered to identify very small genomic losses covered by only 2-9 probes and occurring in at least 5% of the cases (mMCRs). mMCRs having 100% overlap with known copy number variations were discarded.

RESULTS:

mMCRs occurred in 75 known genes. The most commonly affected genes were CDC73 (cell division 73; 63% of the cases, 3 probes), RREB1 (ras responsive element binding protein 1; 60%, 5 probes), JAK2 (47%, 8 probes), CCDC88A (AKT-phosphorylation enhancer,; 47%, 3 probes), AKT3 (43%, 4 probes). Other affected genes at a lower frequency were PIK3CA (26%), EGFR (25%), XRCC4 (18%), JAK1 (18%), PTPRK (15%), RB1 (14%), ERBB2 (10%), PDGFRA (8%), FHIT (7%). A functional analysis performed with DAVID 2008 (http://david.abcc.ncifcrf.gov/) identified the terms “anti-oncogene” and “tyrosine-protein kinase” and five KEGG (http://www.genome.jp/kegg/) pathways (“prostate cancer”, “non-small cell lung cancer”, “pancreatic cancer”, “endometrial” cancer”) as enriched among the 75 genes with a statistically significant p-value <0.05 after Benjamini multiple test correction. Besides tumor suppressor genes such as RB1 and FHIT, very interestingly, many of the genes appeared to code for kinases and for oncogenes. The mMCRs occurred in intronic regions, and apparently targeted highly conserved regions. These regions might represent regulatory loci and their loss may cause gene activation. Validation of selected genes is on-going.

CONCLUSIONS:

The application of high resolution arrays on a large series of CLL samples has shown frequent small interstitial deletions targeting a discrete number of genes, highly enriched for transcripts coding for kinases. A potential mechanism of action might be the loss of regulatory regions determining gene activation. Once validated, the current data would provide the basis to explore the rationale for the use of kinase inhibitors in the treatment of CLL.

Disclosures:

No relevant conflicts of interest to declare.
Ikzf1 Haploinsufficiency Contributes to the Pathogenesis of BCR-ABL1 Positive Acute Lymphoblastic Leukemia.
Authors: Collins-Underwood Racquel , Miller Christopher B , Downing James R. , Mullighan Charles G.
Published: October 2019

Abstract 678

Expression of the constitutively active tyrosine kinase BCR-ABL1 is the hallmark of two diseases with distinct pathologic and clinical features: chronic myeloid leukemia (CML), an expansion of relatively mature granulocytes that typically responds well to kinase inhibition, and pre-B cell acute lymphoblastic leukemia (ALL), an aggressive malignancy of lymphoid progenitors that has a dismal prognosis. The basis for this dichotomy has been poorly understood. Recent studies profiling genome-wide DNA copy number alterations in CML and ALL have identified a near-obligate deletion of IKZF1 (encoding the lymphoid transcription factor IKAROS) in de novo BCR-ABL1 positive ALL, and at the progression of CML to lymphoid blast crisis, suggesting that perturbation of IKAROS activity is a key event in the pathogenesis of BCR-ABL1 lymphoid leukemia. To test this hypothesis, we have examined the effect of Ikaros haploinsufficiency in a retroviral bone marrow transplant model of murine BCR-ABL1 B-progenitor ALL. Unmanipulated marrow from either Ikzf1 wild type mice or mice heterozygous for an Ikzf1 null allele was transduced with MSCV bicistronic retrovirus expressing GFP and p185 BCR-ABL1 and transplanted into lethally irradiated C57BL/6 recipients. Recipients of MSCV-GFP-p185 BCR-ABL1-transduced Ikzf1+/− marrow developed leukemia with markedly increased penetrance and reduced latency in comparison to recipients of wild-type marrow (46 vs. 122 d, P<0.0001). The resulting leukemias resulted in bone marrow replacement by lymphoblasts, pancytopenia, extramedullary infiltration, and frequent clinical and histologic central nervous system involvement. The leukemias were of pre-B immunophenotype and uniformly transplantable into sublethally irradiated secondary recipients. Importantly, detailed enumeration of lymphoid subsets in Ikzf+/+ and +/− marrow did not demonstrate an expansion of lymphoid progenitors in Ikzf1+/− marrow, suggesting that the observed increased penetrance of B-ALL in Ikzf1+/− animals was not simply due to the expansion of targets of BCR-ABL1 transformation. Thus, Ikaros haploinsufficiency cooperates with BCR-ABL1 to induce an aggressive pre-B leukemia with clinical and pathologic features recapitulating human BCR-ABL1 ALL. These results support genomic analyses of human leukemia suggesting that IKAROS loss is important in the pathogenesis of this disease. Experiments evaluating the interaction of Ikzf1 loss with other targets of genomic alteration in BCR-ABL1 ALL (e.g. CDKN2A/B, PAX5), and the role of dominant negative isoforms of Ikaros in leukemogenesis will be of interest.

Disclosures:

No relevant conflicts of interest to declare.
Proteasome Inhibitors Decrease AAV2 Capsid-Derived Peptide Epitope Presentation On MHC Class I Following Transduction.
Authors: Finn Jonathan D , Hui Daniel J , Harre Downey , Dunn Danielle , Mingozzi Federico , Zhou Shangzhen , High Katherine A.
Published: October 2019

Abstract 695

Adeno-associated viral (AAV) vectors are one of the most extensively studied and highly used vector platforms for gene therapy applications. We have recently provided evidence for AAV capsid-derived antigen presentation through MHC class I on the surface of AAV-transduced cells, supporting the hypothesis that in the first clinical trial using AAV to treat Hemophilia B, AAV capsid proteins were presented on the surface of transduced hepatocytes, resulting in clearance by antigen-specific CD8+ T cells and consequent loss of therapeutic transgene expression. Proteasome inhibitors are small molecule compounds that are able to specifically inhibit the activity of the proteasome, resulting in a buildup of ubiquitinated proteins, increased intracellular reactive oxygen species, and a general decrease in presentation of MHCI-peptide complexes. It has previously been shown that proteasome inhibitors can have a dramatic effect on AAV transduction in vitro and in vivo. Here we describe using the FDA approved proteasome inhibitor, bortezomib, to decrease capsid antigen presentation on hepatocytes in vitro, while at the same time, enhancing gene expression in vivo. Using an AAV capsid specific T cell reporter line to analyze effects of proteasome inhibitor on antigen presentation, we demonstrated capsid antigen presentation at low MOI's, as well as inhibition of antigen presentation at clinically relevant levels of bortezomib. We also demonstrate that bortezomib can enhance FIX expression from an AAV2 vector in C57Bl/6 mice, however does not appear to enhance expression of AAV8. Based on the data presented here, it appears as if future studies using proteasome inhibitors in large animal models may be warranted. A pharmacological agent that can enhance AAV transduction, decrease T-cell activation/proliferation, and decrease antigen presentation would be a promising solution to many of the obstacles to successful translation of AAV-mediated, liver-directed gene transfer to the clinic.

Disclosures:

No relevant conflicts of interest to declare.