bluuebunny/crossref_metadata_2025 huggingface dataset:
Gene Therapy for Beta-Thalassemia: Preclinical Studies on Human Cells.
Authors: Ferrari Giuliana , Roselli Emanuela , Tiboni Francesca , Biral Erica , Marktel Sarah , Antoniou Michael , Andreani Marco , Lucarelli Guido , Cesari Rossano , Miccio Annarita
Published: October 2019

Gene therapy for beta-thalassemia is based on the transplantation of genetically-modified autologous hematopoietic stem cells (HSC) into patients affected by the severe form of disease. The genetic treatment of the hemoglobinopathies poses the general challenge of efficient level of gene transfer into HSC and high and persistent transgene expression, in the differentiated progeny of a genetically modified stem cell. The validation of a gene therapy approach to thalassemia requires to obtain results of gene correction in a broad number of patients’ cells, since different molecular defects in the beta-globin gene lead to the clinical phenotype. The heterogeneity in the molecular defects and in the proportion of alpha and non-alpha (beta, gamma and delta) chains will represent a key element to set a threshold in the amount of vector-derived beta-chain required to correct a thalassemic phenotype. Additionally, the impact of some biological parameters, such as the degree of BM erythroid hyperplasia, the BM subpopulations proportion and the apoptotic index, on the successful correction of thalassemic phenotype needs to be studied in the perspective of clinical translation. In order to address these issues, we collected samples from BM aspirates and isolated CD34+ cells from 25 beta+ and beta0 thalassemic patients, characterized by different genotypes and biochemical profiles of globin chains synthesis. A novel, erythroid specific LV expressing human beta-globin from a minimal promoter enhanced by only 2 LCR elements (HS2 and HS3) was used to transduce BM derived CD34+ cells at high efficiency (>80%). The efficacy of the beta-globin LV in correcting the human thalassemic phenotype was tested in an in vitro model of erythropoiesis and in the human-mouse hematological chimera. Upon transduction, normal level of HbA expression was achieved in erythroblastic cultures and BFU-E, associated with a progression towards erythroid maturation, which was impaired in mock-transduced thalassemic cells. Molecular analysis showed proviral integrity, with no detectable rearrangements and an average proviral copy number of 2.4. Analysis of specific globin chains proportion and contribution to phenotype correction in the context of different genotypes is under evaluation.
Clinical Significance of Reticulocyte Hemoglobin as a Diagnostic Tool for Evaluation of Anemia in Veterans.
Authors: Nair Bijay , Spencer Horace J. , Raval Girindra G. , Mehta Paulette
Published: October 2019

Reticulocyte hemoglobin content (CHr), is a reliable marker of iron function, rather than iron stores alone, especially in patients with renal failure. Whether it is as effective in patients with hematologic and oncologic disease is not known. We therefore evaluated the usefulness of CHr for diagnosis of different types of anemia among the Veterans at the CAVHS (Central Arkansas Veteran Healthcare System), Little Rock. We evaluated all patients with anemia seen in the Hematology/Oncology clinic at CAVHS from 7/1/2004 through 6/30/2006. We obtained demographics of patients and laboratory data including CBC, reticulocyte count, CHr, serum iron, total iron binding capacity (TIBC), iron saturation, ferritin, vitamin b12 and folate from the CPRS electronic database. Using STATA program, we analyzed the relation between CHr, RBC indices and other iron storage parameters. A total of 183 patients were evaluated. Patients had mean age 69, ranging from 35 to 98 years. The majority were men with only 9 women. Patients were whites (75%), African Americans (17%), or others (7%). Mean CHr was 32 pg with range from 18.7 to 50.4 pg (normal value 24 to 36 pg). High CHr (>36 pg) was present in 24 patients (13.1 %). High CHr was associated with high MCV in all cases and included patients with vitamin b12 deficiency, myelodysplasia, myelofibrosis, large granular lymphocyte leukemia, chronic lymphocytic leukemia, autoimmune hemolytic anemia, HIV infection on HART as well as those who had received total body irradiation or were taking methotrexate, or azathioprine. Low CHr (<24pg) was present in 8 patients. All of them had low MCV and had iron deficiency anemia, including one with ferritin over 1000ug/ml, presumably high as an acute phase reactant since iron saturation was only 6%. This patient had active head and neck cancer and had received chemotherapy and radiotherapy. CHr did not correlate with other parameters of iron such as ferritin (r=0.14), iron saturation (r=0.57) or TIBC (r= −0.16) in the pooled analysis of all anemias. CHr did correlate well with RBC indices MCV (r=0.86) and MCH (r=0.89) but not so well with MCHC (r= 0.62).

In summary, Reticulocyte hemoglobin content is a good marker of iron function and may be superior to ferritin since it does not change with acute conditions. However it correlates well with MCV and MCH, and therefore does not appear to provide significant independent diagnostic advantage over MCV. Since MCV is readily available and is inexpensive, CHr offers relative little advantage for routine evaluation of anemia.
Dexamethasone+Thalidomide (Dex/Thal) Compared to VAD as a Pre-Transplant Treatment in Newly Diagnosed Multiple Myeloma (MM): A Randomized Trial.
Authors: Macro Margaret , Divine Marine , Uzunhan Yurdagul , Jaccard Arnaud , Bouscary Didier , Leblond Veronique , Janvier Maud , Genet Philippe , Castaigne Sylvie , Royer Bruno , Allard Christian , Chevret Sylvie , Fermand Jean-Paul
Published: October 2019

Since 2003, 204 patients (pts) younger than 66 yrs with untreated active MM were randomly assigned to receive a 4 month treatment with Dex/Thal (n=100) or with a VAD-like regimen (n=104). Then, all pts were intended to proceed to peripheral blood stem cell (PBSC) mobilization using cytoxan 4 g/m2 + G-CSF 5 μg/kg and to receive high dose therapy (HDT) with melphalan 200 mg/m2 and autologous PBSC support. Pts in the Dex/Thal arm received orally Thal for 3 mths at a fixed dose of 200 mg/d combined with Dex, 40 mg/d for 4 days every other wk for 2 mths and then monthly for 2 mths. Pts in the VAD arm received the same Dex regimen plus 3 courses of 4 days continuous infusions of Doxorubicin (9 mg/m2/d for 4 days) and Vincristine (0.4 mg/d for 4 days), 4 wk apart. Anticoagulation prophylaxis wasn’t systematically given.

Main characteristics of pts included in both arms were similar, including age (mean 55.1 yr in the Dex/Thal arm vs 56.4 yr in the VAD arm), MM stage (stage III 78% vs 85%), isotype (IgA: 21% vs 32%, p=.55), serumβ2m (mean 5.2 mg/L vs 3.9 mg/L, p=.24) serum creatinine (≤300 μmol/L in all pts) and albumin levels. Data were analyzed in an intent to treat basis. Primary objective was the achievement of at least a very good partial response (VGPR), defined as a decrease in serum and urine monoclonal immunoglobulin by 90% or greater.

In both arms, 91% of pts proceeded to PBSC mobilization, PBSC harvests were similarly successful and 83% of pts received HDT and autotransplant. Before PBSC collection, 24.7% and 7.3% of pts were in VGPR in the Dex/Thal arm and in the VAD arm, respectively (p=.0027). Before HDT, VGPR rate was 34.7% in the Dex/Thal arm as compared to 12.6% in the VAD arm (p=.002). At 6 mths post transplant, the benefit of Dex/Thal wasn’t further observed with VGPR rates of 44.4% in the Dex/Thal arm and 41.7% in the VAD arm (p=.87). Six pts died in the Thal/Dex arm (including 4 early deaths, within 4 mths post randomization) as compared to 9 pts in the VAD arm (including 3 early deaths).

Venous thrombosis or pulmonary embolism were recorded in 22.8% of pts in the Dex/Thal arm and in 7.5% in the VAD arm (p=.004). A symptomatic peripheral neuropathy was observed in 17.4 % and 12.9% of pts, respectively (p=.42). Otherwise, toxicity profile were similar in both groups. Before PBSC mobilization, mean duration of hospitalization, whatever the cause, was 8.3 days in the Dex/Thal arm as compared to 20 days in the VAD arm (p=.0001).

This randomized study confirms that oral Dex/Thal is an effective first line treatment for symptomatic MM, and supports its use as an induction regimen which could be preferred to infusional VAD in candidates for HDT.
Memory B Cells Protect from CMV Infection in the Absence of T Cells.
Authors: Winkler Thomas H. , Weisel Florian , Klenovsek Karin , Mach Michael
Published: October 2019

Infections with cytomegalovirus are still a major clinical problem in immunosuppressed patients e.g. after bone marrow or stem cell transplantation. To prevent clinical overt disease resulting from disseminated virus infection, immunoprophylaxis and/or -therapy are considered a major goal. The humoral immune response contributes to immune protection against CMV by providing neutralizing antibodies. However, in the early phase after transplantation a primary immune response is not possible. Humoral anti-CMV immune effector functions can only be provided by memory B cells. The activation requirements for resting memory B cells are unclear. Using non-infectious hCMV particles in mice we have recently shown that activation of virus-specific memory B cells to secrete IgG is independent of cognate or bystander T cell help. To analyze whether transfer of memory B cells into immunodeficient mice can protect from lethal infection we switched to an infectious animal model using mCMV. When memory B cells from mCMV-infected mice were adoptively transferred into RAG-1−/− mice, a strong IgG anti-mCMV titer developed within 4–6 days after infection with mCMV. Virus dissemination and subsequent disease was inhibited. A 100–1000 fold decrease of virus titers and a 1.000–10.000 fold decrease of viral DNA load in spleen and lung was observed in mice that received mCMV specific memory B cells. Even in an established mCMV infection virus dissemination and subsequent disease could be prevented by means of adoptive memory B cell transfer. In further experiments we also used a virus mutant that cannot be controlled by NK cells in C57Bl/6 mice. Even in this experimental system we could demonstrate that adoptive transfer of memory B cells in the absence CD4 and CD8 cells is sufficient to protect from viral dissemination and rapid lethality. Our results show that memory B cells can mediate protection against mCMV in the absence of cognate or bystander T cell help. Similar regimens might be a therapeutic option for CMV reactivation after bone marrow transplantation in patients.
Cost-Effectiveness of Rituximab Post CVP Induction for the Treatment of Low-Grade NHL.
Authors: Hornberger John C. , Reyes Carolina , Shewade Ashwini , Loecke David , Valente Nancy
Published: October 2019

Background: Extended Rituximab (R) therapy after CVP (cyclophosphamide, vincristine, prednisone) induction in the first-line treatment of low-grade, CD20-positive, B-cell NHL significantly prolongs progression-free survival (PFS). A societal cost-effectiveness analysis is performed to estimate projected lifetime clinical and economic implications of this treatment.

Methods: A Markov model with 3 health states (progression-free, post-progression, and death) was developed to estimate the direct medical costs and quality-adjusted life-years (QALYs) for a representative patient cohort with low-grade, CD20-positive, B-cell NHL. Kaplan-Meier estimates of PFS and overall survival (OS) up to 4 years are obtained from the ECOG/CALGB intergroup E1496 trial. After 4 years, transition rates are assumed to be the same in both arms. Costs include drug and administration costs, adverse events, treatment of relapses, and end-of-life costs. Incremental costs associated with R are based on Medicare reimbursement rates and published drug prices. Utilities are derived from the literature and a 3% discount rate is employed.

Results: Projected mean OS is 1.2-yrs longer for patients assigned to extended R therapy versus observation alone (15.2 v 14.0 yrs). Extended R therapy is estimated to cost $42,822 on average, with an expected gain of 0.85 years of quality-adjusted survival. Over a lifetime, the cost per QALY gained is $57,515. Sensitivity analyses performed on all variables contributing to the model results showed that utility for follicular NHL patients (range: 0.68–1) and number of R infusions (range: 8–17) most influence the incremental cost-effectiveness ratio (ICER).

Conclusion: The ICER of extended R dosing following CVP induction remains well below the threshold considered acceptable for oncology in the United States. Additional analysis of long term follow-up data will be useful for further evaluation of clinical and economic implications of this treatment for low-grade, CD20-positive, B-cell NHL patients.
Pre Transplantation MGUS and Transformation to Multiple Myeloma in Kidney Transplantation: A Single Center Experience.
Authors: Naina Harris V.K. , Kyle Robert , Habermann Thomas M. , Harris Samar , Cosio Fernando G. , Stegall Mark D. , Leung Nelson
Published: October 2019

Background: Monoclonal gammopathy of undetermined significance (MGUS) is reported in 3 to 5 percent of population, with the prevalence increasing with advancing age. Patients with MGUS are at increased risk for progression to multiple myeloma or other plasma cell dyscrasias. There is a paucity of information on clinical outcomes of patients with MGUS undergoing renal transplantation. A retrospective study was performed to determine wether MGUS is a contraindication to renal transplantation.

Methods: Data was collected from both the kidney transplant and MGUS database. The diagnosis of MGUS was made on the basis of either serum protein electrophoresis (SPEP) or immunofixation after excluding multiple myeloma, amyloidosis and monoclonal immunoglobulin deposition disease.

Results: Between 1977 and 2004, 3518 patients underwent kidney transplantation of whom 23 patients had a preexisting monoclonal gammopathy of undetermined significance (MGUS). Fourteen (61%) of these patients were males. The median age at the time of transplant was 59 ±12 years. Ten patients (43.5%) had IgG Kappa (GK), 7 (30.4%) had IgG Lambda (GL), 2 (8.7%) had IgA Lambda (AL), 1 (4.3%) had IgA Kappa (AK), 2 (8.7%) had IgM Lambda (ML). One patient had a biclonal gammopathy GL and ML. Patients were monitored with either SPEP or immunofixation for median duration of 1542 days after transplantation. Thirteen patients had either no change or stable monoclonal protein, 6 had a decrease in their paraprotein level. Two patients had a mild increase in their paraprotein. Two patients with GK developed into biclonal gammopathy (GK and AK). The median follow up of this cohort after the renal transplant was 1783 days. Twelve (52%) patients remained alive at the time of the study. A patient with GK prior to the transplant who underwent kidney transplantation twice developed a biclonal gammopathy and was found to have increased plasma cells (20%) in bone marrow after 14 years. On follow up for 6 years, his M-protein remained stable. Another patient was found to have 17% plasma cells around the time of kidney transplantation. He had a stable M-protein at follow-up, but underwent a stem cell transplant for recurrent immunotactoid glomerulonephritis. Two (9%) patients developed more than 15% plasma cells in their bone marrow with a stable M-protein. None of the patients with a preexisting MGUS evolved into multiple myeloma.

Conclusion: In this small study, the presence of MGUS prior to kidney transplantation did not appear to have increased the incidence of multiple myeloma post transplant. Therefore, MGUS by itself should not be considered as an absolute contraindication for renal transplantation.
Signaling Profile and Anti-Tumor Activity of the Novel HSP90 Inhibitor NVP-AUY922 in Multiple Myeloma.
Authors: Stühmer Thorsten , Zöllinger Angela , Siegmund Daniela , Chatterjee Manik , Grella Evelyn , Knop Stefan , Kortüm Martin , Unzicker Christian , Jensen Michael R. , Quadt Cornelia , Chène Patrick , Schoepfer Joseph , Garcia-Echeverria Carlos , Einsele Herrmann , Wajant Harald , Bargou Ralf C.
Published: October 2019

We have recently shown that Hsp90 is overexpressed in multiple myeloma (MM) and critically contributes to tumor cell survival. Pharmacologic blockade of Hsp90 has consistently been shown to induce MM cell death. However, most data have been obtained with MM cell lines whereas knowledge about the molecular effects of pharmacologic Hsp90 blockade in primary tumor cells is lacking. Furthermore, these investigations have so far exclusively relied on geldanamycin derivatives. Here, we analyzed the anti-tumor effects of a novel diarylisoxazole-based Hsp90 inhibitor (NVP-AUY922) in a large set of primary MM samples and in MM cell lines. The majority of cell lines (n = 8), as well as most primary samples (n = 20), displayed profound apoptotic responses and steep dose-effect curves with EC50 values in the range of 5–15 nM and EC90 values between 8 and 25 nM. This effect was not attenuated in coculture with cells from the bone marrow microenvironment. Some cell lines and about a quarter of primary MM samples displayed greater resilience to drug treatment, with EC50 values but not EC90 values reached at concentrations up to 50 nM. Sensitivity of MM cells to the Hsp90 inhibitor was not correlated with TP53 mutation or Hsp70 induction levels. Western analyses of MM cell lines and flow cytometric analyses of antibody-stained Hsp90 client proteins in primary tumor cells showed that NVP-AUY922-treatment entailed molecular effects and pharmacodynamic properties consistent with abrogation of Hsp90 function. Consequently, downregulation of multiple signaling and survival pathways was detectable through, for example, decreases in the phosphorylated (activated) forms of extracellular signal-regulated kinase (ERK) 1 and 2, signal transducer and activator of transcription (STAT) 3 and glycogen synthase kinase-3beta. All samples treated displayed strong upregulation of Hsp70. Importantly, peripheral blood mononuclear cells as well as primary bone marrow stromal cells were much less affected by high (50–100 nM) concentrations of NVP-AUY922, showing that a therapeutic window might be established for the treatment of multiple myeloma. Taken together, NVP-AUY922 could be a promising new drug for the treatment of a majority of myeloma patients.
A Prospective Study of Voriconazole Therapeutic Drug Monitoring (TDM) in Patients with Acute Leukemia.
Authors: Seki Jack , Brandwein Joseph , Ward Richard , Paul Narinder , Russell Heather , Kumar Deepali
Published: October 2019

Abstract 1018

Poster Board I-40

Background:

Voriconazole (VOR) is an expanded spectrum azole that is often used to treat suspected or proven invasive aspergillosis (IA). VOR plasma drug levels are not routinely monitored and their correlation with clinical response and toxicity is unclear. We sought to clarify the kinetics of levels in patients started on VOR and their relationship with CYP2C19 genotype and clinical outcomes.

Methods:

We prospectively enrolled inpatients undergoing induction or consolidation chemotherapy for AML who were able to take oral medications and started on VOR for suspected invasive aspergillosis (IA). All patients received an initial intravenous loading dose of 6mg/kg bid for two doses, then 200mg bid. VOR dose could be increased at the physician's discretion. Trough VOR levels (μg/mL) were monitored at days 0,2,4,6 and performed using HPLC. Patients also underwent genotyping for CYP2C19. Clinical and radiologic responses were graded independently by two physicians. Hepatotoxicity was correlated to peak VOR levels.

Results:

We enrolled 33 patients of which 60.6% were male and 39.4% female. Mean age was 60.2±15.4 yrs. The distribution of peak VOR levels was as follows: <2 μg/mL (n=6); 2-6 μg/mL (n=22); >6 μg/mL (n=5). Mild hepatotoxicity (liver enzymes greater than 2x baseline) was seen in 6/22 (27.2%) of patients with levels between 2-6 μg/mL and 1/5 (20%) of patients with levels >6 μg/mL. No hepatotoxicity was noted in those with peak VOR <2 μg/mL. No significant correlation was seen in mg/kg dose of VOR and levels. Of the 33 patients, 19 (57.6%) met the EORTC/MSG criteria for possible, probable, or proven invasive fungal disease. Of these 9 (47.4%) patients had a response to therapy. Mean peak VOR levels in responders vs. nonresponders was 3.78 vs. 3.12 (p=0.63). Response to therapy was 2/2 (100%) if levels were <2 μg/mL, 6/14 (42.9%) if levels were 2-6 μg/mL, and 1/3 (33%) if levels were >6 μg/mL. Breakthrough infections occurred in 5/33 (15.2%) patients given VOR, all of whom had peak levels between 2-6 μg/mL. Genotyping was performed on 11/33 (33.3%) patients and included slow metabolizers (n=5; median peak VOR 3.0), rapid metabolizers (n=5; median peak VOR 3.0) and undetermined (n=1). Genotype did not correlate with clinical outcomes.

Conclusions:

The majority of patients attained voriconazole levels > 2 μg/mL by day 6 of treatment. Voriconazole levels do not appear to predict response to therapy or breakthrough infection although mild hepatotoxicity can occur if levels are >2 μg/mL.

Disclosures:

Seki: Astellas: Honoraria; Pfizer: Honoraria; Merck: Consultancy, Honoraria, Research Funding. Kumar:Merck: Honoraria; Pfizer: Honoraria.
International Normalized Ratio (INR) Determined by Plasma-Based Methods Versus Point-of-Care Instruments in Patients with Antiphospholipid Antibody Syndrome.
Authors: Moll Stephan , McKnight Lauren , Deal Allison
Published: October 2019

Abstract 1067

Poster Board I-89

Background:

In some patients with antiphospholipid antibody (APLA) syndrome treated with warfarin, the International Normalized Ratios (INRs) obtained from plasma, as well as from full blood via fingerstick by older point-of-care (POC) devices, are unreliable. Reliability of newer POC devices in APLA syndrome patients is unknown and was investigated in the present study.

Methods:

60 patients on stable warfarin were enrolled: 30 with APLA syndrome, 30 without. INRs were determined using 4 POCs: CoaguChekXS®, ProTime®, the investigational ProTime®, and INRatio2®. Plasma from phlebotomy was tested for: PT, factor II activity, chromogenic factor X, anticardiolipin and anti-beta-2-glycoprotein I antibodies, lupus anticoagulant. Analysis: Data from the venipuncture INRs from the patients without APLA are used to estimate the relationship between INR and Factor II level and determine the therapeutic range of Factor II. Using the same therapeutic INR range, for each individual POC test the proportion of patients whose clinical decision based on the POC instrument is correct is estimated, along with 95% confidence intervals. This is done for patients with and without APLA. These two proportions are compared between the two groups using Fisher's Exact Tests; nominal p-values are reported. A similar analysis is done using the Factor X test as the gold standard.

Results:

Data collection has been completed; analysis of results is pending and will be available before December 2009.

Conclusions:

Data analysis will show whether the presently available POC devices give reliable INR results in patients with APLA syndrome on warfarin and can be reliably used to monitor warfarin therapy.

Disclosures:

Moll: Hemosense: research support; Roche: reserarch support; ITC: Consultancy, research support. McKnight:ITC: Honoraria. Deal:ITC: Consultancy.
Allogeneic Hematopoietic Cell Transplantation (allo-HCT) Can Induce Durable Remission in Heavily Pretreated Relapsed Hodgkin Lymphoma. (HL).
Authors: Chen Robert , Palmer Joycelynne , Popplewell Leslie , Shen Jessica , Smith Eileen , Delioukina Maria , Kogut Neil , Rosenthal Joseph , Forman Stephen J , Nademanee Auayporn P.
Published: October 2019

Abstract 1192

Poster Board I-214

Background:

Even though Hodgkin lymphoma (HL) is a curable disease, about, 20-30% patients are either refractory to induction chemotherapy or relapse post treatment. High dose chemotherapy and autologous HCT has been shown to be an effective salvage therapy for patients with relapsed HL. However, relapse continues to occur after auto-HCT, especially in patients with chemoresistant or poor-risk features at relapse. The prognosis of these patients is poor with limited options of treatment. Although allo-HCT offers both cytoreduction and potential graft-versus-tumor effect, its use in relapsed HL has been limited by non-relapse mortality (NRM) and patient co-morbidities induced by numerous prior treatments. To examine the potential impact of allo-HCT on survival and disease outcomes, we performed retrospective analysis of allo-HCT in relapsed/refractory HL to determine if allo-HCT can induce long-term remission in heavily pretreated relapsed HL.

Results:

Between January 2003 and December 2008, 29 patients with relapsed HL underwent allo-HCT at City of Hope National Medical Center. The median age was 37 (range: 14-63). 20 (69%) patients were chemosensitive at time of allo-HCT. 17 (59%) patients had prior auto-HCT. 16 (55%) patients received matched siblings and 13 (45%) received unrelated donor cells. 20 (69%) patients had prior radiation treatments. The median number of prior regimens was 5 (range: 2-8). 23 (79%) patients underwent a non-myeloablative conditioning regimen while 6 (21%) patients had a myeloablative regimen. 14 (48%) patients received Tacrolimus/Sirolimus as graft versus host disease prophylaxis and 15 (52%) patients received a combination of Cellcept/CsA, Cellcept/CsA/MTX, Tacrolimus/MTX, or Tacrolimus/Sirolimus/MTX.

With a median follow up of 31.9 months (range: 9.7-69.1) for surviving patients, the results show:

Conclusion:

Allogeneic hematopoietic cell transplantation in heavily pretreated relapsed Hodgkin's lymphoma is feasible, tolerable, and can induce durable clinical remissions.

Disclosures:

No relevant conflicts of interest to declare.
CD137/4-1BB Is Inducibly Expressed On CLL B Cells through CD40 Signaling.
Authors: Fukuda Tetsuya , Nakaima Yukana , Kasubata Aya , Watanabe Ken , Koyama Takatoshi , Rassenti Laura Z. , Kipps Thomas J. , Miura Osamu
Published: October 2019

Abstract 1266

Poster Board I-288

CD137/4-1BB, a TNFR family member is expressed on activated T cells as a co-stimulator. It transduces the signal for cell survival and differentiation and plays a crucial role in CD8 cytotoxic T cells. Recently, an increasing number of reports have indicated its important role on tumor immunity, and the studies of immunotherapy targeting CD137 are on going. Therefore, it is important to know the expression of CD137 and CD137L on malignant cells for the establishment of immunotherapy. As a first step, we examined CD137 expression on PBMCs from healthy donors and CLL patients. When PBMCs from healthy donors were stimulated with PMA and ionomycin, CD137 expression was induced not only on T cells but also on activated B cells. However, when PBMCs from CLL patients were stimulated in the same way, we could not detect CD137 induction on CLL B cells. Since more than 90% of lymphocytes in the patients were CLL B cells, it is conceivable that activated T cells were required to induce CD137 on B cells. To test this hypothesis we next co-cultured CLL cells with T cells activated with anti-CD3/CD28 antibody-coated beads. In this co-culture CD137 was induced on CLL B cells, and this induction was diminished by anti-CD154 blocking antibody. Furthermore, CD137 was inducibly expressed on CLL B cells after co-culture with HeLa cells transfected with CD154 gene. The induction of CD137 mRNA was also clearly detected by RT-PCR after this stimulation. This CD137 induction was more significantly observed on CLL B cells (n=14, MFIR 11.5±6.9) as compared with B cells from healthy donors (n=4, MFIR 3.7±0.6, p=0.001) or non-CLL B cell malignancies (n=9, MFIR 4.2±3.43, p=0.003). Stimulation of CD137 expressed on BJAB transfectants by co-culture with CD137L-transfected CHO cells induced a conspicuous nuclear translocation of p52, a non-canonical NF-κB factor. In agreement with this activation, the expression of survival factor BCL-XL was upregulated. Consequently, the CD137 signal augmented the survival of CD154 stimulated CLL B cells in vitro. CD40 ligation can induce anti-CLL immunity and reduce CLL cells clinically. These data suggest that the inducibly expressed CD137 may diminish the effectiveness. It is possible that the fine adjustment of these co-stimulators can lead more effective immunotherapy.

Disclosures

No relevant conflicts of interest to declare.
Inhibition of MALT1 Protease Activity Is Selectively Toxic for Activated B CellÃ,Â–Like Diffuse Large B Cell Lymphoma Cells.
Authors: Ferch Uta , Kloo Bernhard , Gewies Andreas , Pfaender Vera , Düwel Michael , Peschel Christian , Krappmann Daniel , Ruland Jürgen
Published: October 2019

Abstract 1271

Poster Board I-293

Diffuse large B cell lymphoma (DLBCL) is the most common type of lymphoma in humans. The aggressive activated B cell-like (ABC) subtype of DLBCL is characterized by constitutive NF-κB activity and requires signals from CARD11, BCL10 and the paracaspase MALT1 for survival. CARD11, BCL10 and MALT1 are scaffold proteins that normally associate upon antigen receptor ligation. Signal-induced CARD11/BCL10/MALT1 (CBM) complexes couple upstream events to IKK/NF-κB activation. MALT1 possesses in addition a recently recognized proteolytic activity that cleaves and inactivates the negative NF-κB regulator A20 and BCL10 upon antigen receptor ligation. Yet, the relevance of MALT1 proteolytic activity for malignant cell growth is unknown. Here we demonstrate pre-assembled CBM complexes and constitutive proteolysis of the two known MALT1 substrates in ABC-DLBCL but not in germinal center B cell-like (GCB) DLBCL. ABC-DLBCL cell treatment with a MALT1 protease inhibitor blocks A20 and BCL10 cleavage, reduces NF-κB activity and decreases the expression of NF-κB targets genes. Finally, MALT1 paracaspase inhibition results in death and growth retardation selectively in ABC-DLBCL cells. Thus, our results indicate a growth-promoting role for MALT1 paracaspase activity in ABC-DLBCL and suggest that a pharmacological MALT1 protease inhibition could be a promising approach for lymphoma treatment.

Disclosures

No relevant conflicts of interest to declare.
Impact of Treatment Intensity and Factor VIII Products On the Development On High Titre Inhibitors in Children with Severe Hemophilia A: Results of a Non-Concurrent Cohort Study.
Authors: Kurnik Karin , Halimeh Susan , Manner Daniela , Holzhauer Susanne , Ettingshausen Carmen Escuriola , Schobess Rosemarie , Nowak-Gottl Ulrike
Published: October 2019

Abstract 1303

Poster Board I-325

Background

A number of environmental and genetic factors have been identified to influence inhibitor development in children with severe hemophilia A (HA): among them, individual therapeutic regimens and the use of different factor VIII products have been controversially discussed.

Methods

In the present multicenter cohort study we evaluated the impact of i) treatment intensity [median individual dose administered during the first 6 to 8 weeks] and ii) type of factor VIII product [plasma-derived (pd); recombinant-derived (r)] on the risk of high-titre inhibitor development in 150 previously untreated patients with severe hemophilia A (HA) consecutively ascertained between 1982 and 2007 from five German catchment areas. Patients were followed over a period of 200 exposure days from HA onset. Study endpoint was inhibitor-free survival time related to treatment with adjustment for treatment period. Plasma levels of factor VIII were determined by one-stage clotting assays. Inhibitor testing was performed at least monthly to 3 monthly when on therapy using the Bethesda method or its modification [Nijmegen]: The lower detection limit was set at 0.6 Bethesda units [BU]. A peak inhibitor titer of > 5 BU was defined as high responder [HR]. A positive inhibitor testing was stated when an inhibitor was measured at least in two independent follow-up visits. The cumulative inhibitor-free survival was calculated using multivariate Cox regression [hazard ratio (HR) and 95% confidence intervals [CIs]].

Results

During the follow-up period 30 of 150 children (20.0%) developed a high titre inhibitor. In univariate analysis the median dose increase per IU/kgbw significantly increased the hazard towards HR inhibitor development [HR/CI: 1.08/1.05-1.11]. In addition, 14 of 52 children treated with rFVIII concentrates (27%) versus 16 of 98 children (16.3%) treated with pdFVIII concentrates developed HR during the observation period [HR/CI: 1.9/0.9-3.9]. In multivariate analysis treatment intensity adjusted for FVIII products and treatment period were independently associated with the development of clinical meaningful inhibitors [HR/95%CI: 1.08/1.1-1.1].

Conclusion

Data presented here support the hypothesis that clinical meaningful inhibitor development is of multifactorial origin and that treatment intensity plays a major role.

Disclosures

Off Label Use: Enoxaparin (LMWH) is used off-label in children to prevent symptomatic thromboembolism.
Reduced Number of Dense Bodies and Reduced Serotonin Content in Platelets of Patients with Wiskott-Aldrich Syndrome.
Authors: Sokolic Robert A. , Markello Thomas , Krumwiede Marcy , Fischer Roxanne , Garabedian Elizabeth , Gahl William A. , White James G. , Candotti Fabio
Published: October 2019

Abstract 1321

Poster Board I-343

Wiskott-Aldrich syndrome is characterized by immune deficiency and thrombocytopenia. There is also a qualitative platelet defect in Wiskott-Aldrich syndrome, typically presenting on aggregometry as a failure of the second wave of aggregation. The pathophysiology of this defect has not been studied in detail. To investigate this phenomenon, we enumerated dense bodies and measured serotonin content in the platelets of 7 patients with Wiskott-Aldrich syndrome. Patients ranged in age from 14 to 45 years old. Five of 7 patients had had splenectomy. In these patients, the platelet count ranged from 120,000/mcL to 521,000/mcL. Platelet counts were 20,000/mcL and 46,000/mcL in the 2 patients who had not had splenectomy. The latter 2 patients had occasional petechiae, as did 2 of the splenectomized patients. Three patients had no clinical evidence of platelet dysfunction. One of these 3 had tolerated abdominal surgery without untoward bleeding. Blood specimens were taken in the course of routine follow-up, when patients were otherwise well. Platelets were prepaed for electron microscopy by the whole mount technique. For serotonin content, platelets were counted and lysed in distilled water. Serotonin in the lysate was chemically acetylated and then measured by ELISA. Serotonin levels were normalized to ng of serotonin per 10 9 platelets. The average ± standard deviation dense body count in our patient group was 1.1 ± 0.4 dense bodies per platelet (normal count 6-8 dense bodies/platelet). Serotonin content was measured in three patients, who showed 273 ± 103 ng serotonin per 10 9 platelets (normal value 621-1064 ng/10 9 platelets). Our results are the first quantitative analysis of platelet dense body counts and serotonin level in patients with Wiskott-Aldrich syndrome. In addition to further elucidating the pathophysiology of this disease, these data have implications for understanding the role of the Wiskott-Aldrich syndrome protein in the genesis of dense bodies.

Disclosures

No relevant conflicts of interest to declare.
Induction of Potent Anti-Tumor Effects by Original and TCR-Redirected CD4 + Cytotoxic T Cells.
Authors: Spaapen Robbert M , van den Oudenalder Kelly , van Elk Maureen , Aarts Tineke , Guichelaar Teun , Groen Richard WJ , Bloem Andries C , Storm Gert , Martens Anton C , Lokhorst Henk M , Mutis Tuna
Published: October 2019

Abstract 1333

Poster Board I-355

The curative Graft versus Tumor (GvT) effect of allogeneic stem cell transplantation and donor lymphocyte infusions is mainly mediated by donor derived T cells recognizing minor Histocompatibility antigens (mHag) presented by malignant cells. Traditionally, CD8+ cytotoxic T cells (CTLs) are considered as “the” effector cells of these anti-tumor responses, whereas a sole helper role is attributed to CD4+ T cells. CD4+ T cells often possess killer capacities in vitro, raising the possibility that they may also mediate anti-tumor effects without the need for CD8+ T cells. Hence, we here explored the feasibility of adoptive immunotherapy with sole CD4+ CTLs by testing the therapeutic capacity of a mHag-specific, CD4+ CTL (3AB11) in a human GvT model. Rag2−/−γc−/− mice were inoculated with BLI detectable, mHag+ Multiple myeloma cells. Treatment of established tumors with 3AB11 but not with control CD4+ T cells by triple consecutive injections of 30-40×106 cells/day rapidly reduced the medullary growing tumors, illustrating for the first time the feasibility to establish a significant GvT effect by targeting a sole mHag recognized by CD4+ CTLs. The therapy was less effective at a higher tumor load and unsuccessful by a single injection of 20×106 cells, underscoring the critical importance of T cell dose-to-tumor load ratio to establish an efficient anti-tumor effect. In further exploration, tumors were also significantly reduced by treatment with “dual antigen-specific” T cells, which were generated by transduction of the T cell receptor (TCR) of clone 3AB11 into recall antigen (Tetanus Toxoid; TT)-specific T cells. Finally, the in vivo persisting dual-specific T cells could be boosted by administration of TT loaded mHag negative B cells, demonstrating for the first time the feasibility and potential advantages of immunotherapy with TCR-transduced “dual antigen-specific” CD4+ T cells. We conclude that potent GvT effects may be achieved in clinical trials by targeting a sole mHag antigen with original or TCR-redirected CD4+ CTLs.

Disclosures

No relevant conflicts of interest to declare.
Avascular Necrosis in Untreated Patients with Type 1 Gaucher Disease.
Authors: Mistry Pramod K , Deegan Patrick , Vellodi Ashok , Cole J. Alexander , Yeh Michael , Weinreb Neal J.
Published: October 2019

Abstract 1353

Poster Board I-375

Objective

To determine the incidence of avascular necrosis (AVN) in untreated patients with type 1 Gaucher disease (GD1).

Methods

All patients with GD1 enrolled in the ICGG Gaucher Registry as of July 2007 were included in this analysis. All GD1 patients who either never received treatment or eventually went on to receive treatment were identified. Follow-up began on each patient's date of earliest reported assessment in the Registry. Among patients who never received treatment, follow-up continued until the last recorded assessment date in the Registry. For patients who eventually went on to receive treatment, follow-up continued until the date of initiation of therapy. Incidence rates (and Poisson exact 95% confidence intervals) of AVN were determined for both groups of patients. AVN was typically ascertained from X-Ray or MRI results.

Results

As of July 2007, the inclusion criteria were met by 3,497 patients. The incidence rate of AVN among untreated patients was 22.8 per 1,000 person years (95% CI 20.2 to 25.7). Patients with antecedent splenectomy (total or partial) had a higher incidence rate of AVN (46.6 per 1,000 person-years, 95% CI 38.4 to 56.1) compared to patients without a splenectomy (incidence rate 17.0 per 1,000 person-years, 95% CI 14.5 to 19.8). The primary sites where AVN was identified in both groups were the hip and femur.

Conclusion

This is the first epidemiologic analysis to estimate incidence rates of AVN among untreated patients with GD1. Splenectomy appears to be a risk factor for GD1-associated AVN. Based on the incidence results above and presupposing equal risk distribution, without therapeutic intervention, most patients should theoretically experience at least one episode of AVN at some point in their life. However, because the GD1 population is genotypically and phenotypically heterogeneous, further analyses will attempt to identify characteristics that distinguish untreated patients at high risk of developing AVN from those who are less likely to develop this serious complication.

Disclosures

Mistry: Genzyme Corporation: Honoraria, Research Funding. Deegan:Genzyme Corporation: Honoraria. Vellodi:Genzyme Corporation: Honoraria, Speakers Bureau. Cole:Genzyme Corporation: Employment. Yeh:Genzyme Corporation: Employment. Weinreb:Genzyme Corporation: Honoraria, Membership on an entity's Board of Directors or advisory committees.
Characterization of Nur77+ Cells in Murine Peripheral Blood.
Authors: Hubbeling Harper , Beck Thomas , Moran Amy , Hogquist Kristin , Maltzman Jonathan , Cunningham Nicole , Punt Jenni
Published: October 2019

Abstract 1361

Poster Board I-385

The expression of Nur77/NR4A1, one member of the Nur Orphan Nuclear Receptor family, is tightly regulated in many cell types, including myeloid and lymphoid lineages, and can change rapidly in response to a variety of external stimuli. Best known for its role in regulating apoptosis by either transcribing lethal genes or by initiating mitochondrial production of cytochrome c, Nur77 may also play other non-apoptotic roles. In an effort to understand the fate and function of cells that upregulate Nur77, we developed a transgenic Nur77-eGFP reporter mouse. Our analysis of cells in the peripheral blood of these mice reveals two populations of circulating cells that express GFP (Nur77) ‘constitutively’: T lymphocytes and a subset of monocytes. Our preliminary data suggest that the GFP+ T cells, which, interestingly are enriched for regulatory T cells, are recent thymic emigrants that have upregulated Nur77 as a consequence of TCR stimulation during maturation. Our observations also indicate that the GFP+ monocytes define a subpopulation of ‘patrolling’ cells that may ultimately differentiate into splenic dendritic cells. Understanding the identity of these circulating GFP+ cells should provide us with insights into the regulation and function of Nur77 itself. The analysis may also clarify the significance of phenotypic differences among blood cell subtypes by providing additional information about cell origin and activity.

We thank the Arnold and Mabel Beckman Foundation (HH), HHMI (TB), and NSF (#MCB-0744570 (JP and NC)) for their support for this work.

Disclosures

No relevant conflicts of interest to declare.
Meprin αβ Deficiency Results in Accumulation of Peritoneal Macrophages and Hyperactive Pro-Inflammatory Cytokine Response in Mice.
Authors: Jin Hong-Jian , Banerjee Sanjita , Yura Renee , Bao Lei , Bond Judith , Sun Qi
Published: October 2019

Abstract 1364

Poster Board I-386

Meprin metalloproteases are abundantly expressed in kidney and intestinal epithelial cells, and have been implicated in inflammatory bowel disease. Recently, we demonstrated that meprin deficiency compromises homeostasis of monocytes and natural killer cells. However, the role(s) of meprins in the hematological system and the pathological consequences of meprin deficiency have not been fully characterized. We report here that mouse and human monocytes expressed meprins, and the expression of meprin α was down-regulated in monocyte-derived mature macrophages (Mφ). In a mouse model of septic shock induced by peritoneally injected bacterial lipopolysaccharide (LPS), mice carrying germline null mutation of both meprin α and βgenes (meprin double knockout, dKO) developed a more severe form of septic shock, characterized by deeper hypothermia, higher levels of blood nitrate/nitrites and suppressed monocytosis, in comparison to the wild-type mice. In addition, LPS-challenged meprin dKO mice exhibited a greater elevation of plasma IL-12, MCP-1, IL-6 and TNF-α, even though these pro-inflammatory cytokines were at lower levels in the dKO mice than the wild-type mice in the steady state. Furthermore, meprin dKO mice harbored a higher number of peritoneal Mφ than the wild-type mice, and thereby produced more soluble cytokines in response to ex vivo LPS stimulation. These results indicate that meprin deficiency is associated with accumulation of peritoneal Mφ, which may lead to hyperactive cytokine responses to inflammatory stimuli and result in increased susceptibility to LPS-induced septic shock.

Disclosures

No relevant conflicts of interest to declare.
Temporal Trends in the Proportion Cured Among Patients Diagnosed with Acute Myeloid Leukemia in Sweden 1973-2001, a Population-Based Study.
Authors: Derolf Åsa Rangert , Andersson Therese M-L , Lambert Paul C , Kristinsson Sigurdur Y , Eloranta Sandra , Landgren Ola , Dickman Paul W , Bjorkholm Magnus
Published: October 2019

Abstract 1378

Poster Board I-400

Background:

Acute myeloid leukemia (AML) is the most common type of acute leukemia in adults and is rapidly fatal without specific therapy. In a recently published population-based study we showed large differences in 1- and 5-year relative survival in AML patients in Sweden depending on age at diagnosis and year of diagnosis[1]. Here we use an alternative approach to study patient survival that simultaneously estimates the proportion of patients cured from AML and the survival time of those patients that are not cured. The cure proportion provides a better estimate of long-term survival than 5-year survival and is therefore of direct interest to patients and health care professions.

Methods:

We conducted a population-based cohort study including 6,439 patients aged 19-79 diagnosed with AML in Sweden 1973 to 2001. Patients are considered statistically cured when, as a group, their mortality returns the level of a comparable general population. We estimated mixture cure fraction models that provide estimates of both the cure proportion and the distribution of survival times of the “uncured”. Age at diagnosis was classified into four categories 19-40, 41-60, 61-70 and 71-80 and year of diagnosis was modeled using restricted cubic splines.

Results:

During the first years of the study period the cure fraction was less than 5% for all age groups and the median survival time for “uncured” approximately 0.5 years or less. In 2000 the estimated cure proportion was 68% (95% CI 56%-77%) for the youngest age group, 32% (25-39%) for those aged 41-60, 8.4% (3.1-21%) for those aged 61-70 at diagnosis and 4.3% (2.3-8.1%) for the oldest age group. The estimated median survival times for “uncured” were 0.74 (0.43-1.26), 0.71 (0.53-0.97), 0.69 (0.51-0.95) and 0.37 (0.31-0.44) years respectively (Table 1). The improvement in survival manifested as longer survival among the uncured during the early calendar years but increases in the cure proportion were observed during later years.

Conclusion:

There are large differences in the proportion cured between the age groups. In younger patients the cure proportion has increased dramatically, while survival of the “uncured” actually decreased in the last time period. In the older age groups improvement is merely seen within the survival of the “uncured”.

[1] Derolf AR, Kristinsson SY, Andersson TM-L, Landgren O, Dickman PW, Björkholm M. Improved patient survival for acute myeloid leukemia: A population-based study of 9,729 patients diagnosed in Sweden 1973-2005. Blood. 2009 Apr 16;113(16):3666-72.

Disclosures:

No relevant conflicts of interest to declare.