bluuebunny/crossref_metadata_2025 huggingface dataset:
Combined VBM Chemotherapy and Involved Field (IF) Radiotherapy in Early Stage Hodgkin’s Lymphoma (HL): A Single Centre Experience.
Authors: Cocorocchio Emilia , Peccatori Fedro A. , Gigli Federica , Bassi Simona , Nassi Luca , Calabrese Liliana , Travaini Laura , Piperno Gaia , Preda Lorenzo , Pruneri Giancarlo , Liptrott Sarah J. , Alietti Alessandra , Martinelli Giovanni
Published: October 2019

Early stage HL patients (pts) benefit from a short course of chemotherapy combined with IF radiotherapy. In our previous experience six courses of VBM chemotherapy (Stanford regimen) were administered with IF radiotherapy. However, despite good clinical results, the schedule was modified in terms of total chemotherapy cycles delivered, in order to reduce the risk of long term toxicities. PET and CT scan were performed at staging to identify a low risk patients (pts) group. From May 2001 to November 2006, 32 treatment naïve Hodgkin’s lymphoma pts (18 male, 14 female) were treated with four courses of VBM chemotherapy (D1, 8: Vinblastine 6 mg/sm, Bleomycin 10 mg/sm, Methotrexate 30 mg/sm, repeated every 28 days) followed by IF radiotherapy. Median age was 37 years (range 16–73). Main histology was classical HL in 26 pts, while lymphocyte predominant was observed in 6 pts. Pts were all stage I-IIA (no bulky disease), no more than three sites involved, performance status 0–1 and ESR < 50. CT scans were performed before and after chemotherapy and radiotherapy. PET scans were performed before and after chemotherapy in 26 pts, and after radiotherapy if not performed before or in case of positivity after chemotherapy. Chemotherapy was usually well tolerated: three pts experienced G3-4 neutropenia while the main non- haematological toxicities recorded were G3 transaminitis in 3 pts and G1-2 neuroconstipation/abdominal pain in 18 pts. IF radiotherapy was performed one month after the last chemotherapy cycle; median dose administered was 30Gy (range 26–30.4). At the end of chemotherapy 7 pts were PET positive. While one patient developed progressive disease, the other six pts became negative after radiotherapy and are still in complete remission (CR). 29 pts (90%) completed the whole program obtaining a CR. Two pts progressed before radiotherapy, while one patient shifted to an anthracycline containing schedule after two cycles due to stable disease. With a median follow-up of 42 months (range: 7–73), all pts are alive. 1 out of 29 CR pts progressed and is now receiving intensive chemotherapy program with autologous stem cell transplant. In our experience four cycles of VBM chemotherapy combined with IF radiotherapy is an effective program. Furthermore this regimen that doesn’t contain alkylating agents or anthracyclines avoids alopecia and decreases the risk of long term toxicity. PET scan effectively helps to identify low risk early stage HL pts that can take advantage of this therapeutic approach.
Abnormally High Levels of Circulating CD34+CD38−, CD34+CD38+, and CD10+ Hematopoietic Stem and Progenitor Cells in the Peripheral Blood of B-Cell Chronic Lymphocytic Leukemia (B-CLL) Patients Suggest the Presence of Additional Therapeutic Target Cells.
Authors: Christopherson Kent W. , Paganessi Laura A. , Rhoades Darilyn R. , Dillon Hortense M. , Gezer Sefer , Larson Melissa L. , Fung Henry C. , Gregory Stephanie A.
Published: October 2019

Chronic Lymphocytic Leukemia (CLL) is a lymphoproliferative disorder manifested by a clonal expansion of mature long-lived functionally defective lymphocytes that are predominantly of B-cell origin (>95% of cases). CLL cells typically express the pan-B-cell antigens CD19 and CD20 as wells as CD5, an antigen of mature T-cells, which is present in 95% of B-CLL patients and is helpful for diagnosis. The disease has a median survival of 8–10 years. Asymptomatic patients (RAI stages 0–2) are often not treated. RAI stages 3 & 4 are generally treated. Current therapies for CLL included a combination of monoclonal antibodies (anti-CD20 monoclonal antibody, rituximab) with purine analogs (fludarabine or pentostatin) and alkylating agents (cyclophosphamide). Current therapies to manage B-cell CLL are not curative and therefore suggest that the current strategy of targeting therapy against B-cells may leave behind residual disease that contributes to progression or relapse. The importance of more primitive Hematopoietic Stem and Progenitor cell populations in B-CLL patients has not been considered. We hypothesized that circulating cells may be spontaneously mobilized into the periphery of CLL patients and would therefore contribute to the peripheral blood microenvironment. To test this hypothesis we examined existing CLL cell populations from un-treated “watch-and-wait” B-CLL patients as compared to normal volunteer donor peripheral blood cell populations utilizing multivariate flow cytometric analysis. We determined CD34+CD38−, CD34+CD38+, and CD10 levels in normal peripheral blood to be 81.0±14.5, 497.3±78.33, and 83.6±17.5 cells/ml respectively. We report here the presence of abnormally high levels of these stem and progenitor cell populations in B-CLL patients (see table). Although the mechanism behind the release of these cells into the periphery in B-CLL patients is unknown, it is reasonable to propose that they may be contributing to the manifestation of the disease and are therefore potential novel target cell populations for future therapies.

B-CLL Patients WBC β2m CD34+CD38− CD34+CD38+ CD10+ White Blood Cell (WBC) counts are expressed as 1x106 cells/ml. All other counts are expressed as cells/ml 7.93 2.4 2,526 14.6 23,119 11.86 U 3,145 58.2 1,449 12.5 U 307 11.72 1,773 16.9 3.7 6,450 251 3,771 17.24 3.0 28 5.79 8,024 19.37 1.8 4,086 180 8,247 19.9 U 1,790 18.6 3,099 19.92 U 529 34.1 16,039 25.81 1.8 2,937 194 4,617 31.21 2.9 20,753 304 24,340 43.13 1.5 1,577 260.49 281,976 73.98 3.2 1,099 1,767 77,525 105.53 2.2 3,266 220 24,070
2-Chlorodeoxyadenosine (2-CDA) and Dexamethasone Induce Apoptosis in Multiple Myeloma (MM) Via Different Mechanisms.
Authors: Liu Bolin , Fang Zeying , Ma Jian , Dillon Thomas E. , Byers Tim E. , Thor Ann D. , Lee Choon Kee
Published: October 2019

Although 2-CDA has been active on B-lymphocyte derived malignancies, its potential for myeloma growth control has not been fully investigated. In the present study on a pair of MM cell lines, dexamethasone-sensitive (MM1.S) and dexamethasone-resistant (MM1.R), we sought to determine whether 2-CDA can effectively induce apoptosis and growth inhibition in both cell lines and whether there is cross resistance between 2-CDA and dexamethasone in MM1.R cells. Cell proliferation assay (MTS) releaved that 2-CDA significantly inhibited both MM1.S and MM1.R cell growth in a dose dependent manner, with the minimum (10%) and maximum (100%) inhibition concentration of 12.5 nM/L and 500 nM/L for the MM1.S, and 25 nM/L and 500 nM/L for the MM1.R cells, respectively. The IC50 values of 2-CDA in the MM1.S and MM1.R cells were 48 nM/L and 60 nM/L, respectively. No cross resistance was observed between 2-CDA and dexamethasone in the MM1.R cells. On the molecular level, dexamethasone induced PARP and caspase-9 cleavage, and increased the level of p27kip1 only in the MM1.S cells. 2-CDA treatment in both cell lines resulted in DNA fragmentation as well as strong PARP and caspase-9 cleavage, but no significant changes in the levels of P-Akt, P-MARK, p27kip1, E2F1, and cyclin D1, indicating that 2-CDA induces growth inhibition and cell death in MM cell lines likely through mitochondria-dependent apoptotic mechanism. Cell cycle analyses by flow cytometry showed that dexamethasone (5μM/L) treatment increased sub-G1 (apoptosis) cells to 8.1% only in the MM1.S cells, while the majority (87%) of cells were arrested in the G1 phase of cell cycle in 24 hours. In contrast, 2-CDA (0.5μM/L for 24 hours) strongly induced apoptosis in both cell lines (sub-G1 population increased to 19.6% and 22.1% for the MM1.S and MM1.R cells, respectively) without changing their cell cycle profiles. These data suggest that dexamethasone and 2-CDA induce apoptosis in myeloma cells via different mechanisms, which would provide a theoretical basis for combination therapy for MM with these two agents. Furthermore, our results also show that 2-CDA alone is capable of inducing apoptosis in the MM1.R cells, suggesting that 2-CDA may have therapeutic potential for MM patients with a dexamethasone-resistant phenotype.
Defibrotide for the Treatment of Veno-Occlusive Disease Post SCT: Preliminary Results of EU Compassionate Use Program in 332 Patients Confirm Promising Activity and Manageable Toxicity.
Authors: Carreras Enric , Olavarria Eduardo , Neven Benedicte , Maertens Johan , Boelens Jaap J. , Tescione Francesco , Hoyle Margaret , Ferro Laura I. , Iacobelli Massimo , Ruutu Tapani , Niederwieser Dietger
Published: October 2019

Introduction: Hepatic veno-occlusive disease (VOD) is one of the most significant regimen-related toxicities of stem cell transplantation (SCT), which when complicated by multi-organ failure (MOF) has a case fatality rate >90%. Defibrotide (DF) is a polydisperse oligonucleotide, with anti-thrombotic, anti-ischemic and thrombolytic properties, especially on microvasculature. Results in earlier studies of DF therapy for VOD/MOF have consistently shown manageable toxicity with promising complete response (CR) rates and D+100 survival post SCT. We report the results of the largest compassionate use program (CUP) performed to date.

Methods: Physicians in Europe requesting DF for the treatment of pts with VOD obtained the drug on a named patient basis and were requested to collect data by a standardized CRF. Patients (pts) were reviewed to identify the nature and frequency of Suspected Adverse Drug Reactions (SADR). CR and D+100 survival post-SCT were assessed with CR defined as bilirubin <2 mg/dl and resolution of VOD-related end-organ dysfunction. From February 1999 to July 2007, 332 pts (215 men (65%), 117 women (35%), with a median age of 28 years) from 96 European centers received DF (Gentium SpA, Como, Italy). Pts were both adults (208 pts 63%, range: 18–66 yrs) and pediatric (124 pts, 37%, range: 2 months-17 years). 253 pts underwent allo-SCT (77%), and 76 auto-SCT (23%). At entry, all pts had VOD defined by either Baltimore criteria (205, 62%), modified Baltimore criteria [within 35 days of SCT] (n=42, 13%), Seattle criteria (84, 25%) or were biopsy proven (n=1). MOF was present in 113 pts (34%): 90/332 (27%) pts had renal, 53/332 (16%) had respiratory and 15/332 (5%) had CNS dysfunction. Median dose of DF was 37 mg/kg/d (range: 10–120 mg/kg/d). Median duration of DF therapy was 14d. Overall, D+100 survival was 46% (154 pts). CR was achieved in 173 pts (52%), with 142 alive in CR at D+100 (82%). NR was seen in 159 (48%), with only 12 alive at D+100 (8%). In a subset of 73 pts with severe VOD (defined by Baltimore criteria and MOF), survival at D+100 was 34% (25 pts). The overall incidence of attributable SADRs was 8% (n=28) and consisted predominantly of haemorrhages (n=27, 8%), most of which proved reversible.

Conclusions: In this largest study of pts treated with DF to date, no unexpected toxicities were reported and side effects were generally manageable. CR was obtained in 52% with D+100 survival in 46%, including 34% in those pts with severe disease. This is consistent with prior studies and suggests that DF offers a relatively safe and effective treatment for hepatic VOD post SCT.
CEACAM-1 Is Involved in Graft-Versus-Host-Disease in Murine Allogeneic Bone Marrow Transplantation Models.
Authors: Lu Sydney X. , Willis Lucy , Smith Marsinay , Suh David , King Christopher , Bautista Johanne , Chow Melanie , Cabrera-Perez Javier , Hubbard Vanessa , Rotolo Jimmy , Liu Chen , Murphy George , Alpdogan Onder , Blumberg Richard , Holmes Kathryn , Turbide Claire , Beauchemin Nicole , van den Brink Marcel R.M.
Published: October 2019

Carcinoembryonic antigen associated cell adhesion molecule 1 (CEACAM-1) belongs to a family of carcinoembryonic antigen-associated glycoproteins. It is expressed on leukocytes, endothelium, and epithelium. Microarray analysis showed that CEACAM-1 mRNA is increased in the small bowel during gut graft-versus-host-disease (GVHD) after allogeneic bone marrow transplantation (allo-BMT). Using CEACAM-1−/− mice as recipients or sources of donor bone marrow or T cells caused significantly worse GVHD mortality (p<0.05) compared to wildtype (WT) controls. Histopathological analysis of GVHD target organs from CEACAM-1−/− recipients of WT T cells or WT recipients of CEACAM-1−/− T cells revealed increased GVHD of the large bowel (p<0.05) but not liver or small bowel compared to WT control. Alloreactive splenic CD8 CEACAM-1−/− T cells from recipients with GVHD had increased levels of α4β7 integrin compared to WT controls. We also found increased numbers of small bowel intraepithelial lymphocytes and mesenteric lymph node cellularity in CEACAM-1−/− recipients of WT T cells and WT recipients of CEACAM-1−/− T cells, with a corresponding decrease of cellularity in peripheral lymph nodes and the liver. Adoptive transfer of CFSEhi CEACAM-1−/− T cells into WT hosts, or of WT T cells into CEACAM 1−/− hosts revealed more profound activation of T cells in CEACAM-1 deficient settings, shown by increased early CD25 expression and CD62L down-regulation on splenic CFSEdim alloreactive T cells. We found no significant differences in serum levels of TNF or IFNγ, T cell proliferation kinetics upon adoptive transfer, percentages of alloactivated CD4 or CD8 cells, intracellular levels of T-bet or IFNγ, CD8 T cell cytolytic efficiency, percentages of splenic regulatory T cells, or levels of T cell apoptosis in WT recipients of CEACAM-1−/− T cells or CEACAM-1−/− recipients of WT T cells with GVHD as compared with controls. Finally, irradiation of non-transplanted CEACAM-1−/− mice revealed increased radiation sensitivity, shown by earlier and greater lethality and increased small bowel crypt apoptosis, suggesting a role for CEACAM-1 in conditioning-related toxicity and subsequent GVHD amplification. We conclude that CEACAM-1 deficiency on donor T cells or transplant recipients results in increased gut and systemic GVHD due to increased T cell activation and elevated expression of the gut homing integrin α4β7. This suggests that the use of CEACAM-1 agonists could be a novel theraputic strategy for ameliorating acute intestinal and systemic graft-versus-host-disease.
Mutations in the Catalytic Subunit of Class IA PI3K Are Leukemogenic In Vivo.
Authors: Horn Stefan , McCubrey James A. , Stocking Carol , Trumper Lorenz , Basecke Jorg
Published: October 2019

The phosphoinositide 3-kinase (PI3K)-AKT pathway is a relevant signal transduction axis which regulates survival, growth and proliferation of hematopoietic cells through a variety of downstream targets. Constitutive activation of the PI3K-AKT pathway is observed in up to 70% of acute myelogenous leukemia (AML), primarily due to activating mutations in receptor tyrosine kinases (e.g. c-KIT) or RAS and is also present in T-cell-non-Hodgkins-lymphoma, mantle cell lymphoma, diffuse large B-cell lymphoma, Hodgkins Lymphoma and myelodysplastic syndrome (MDS). We analyzed the effect of activating point mutations in the catalytic subunit p110alpha of class IA PI3K (PIK3CA) in hematopoietic cells. We transfected early hematopoietic, IL-3 dependent cells (Ba/F3 cell line) with point mutations in the helical (exon 9, E542K, E545A) and kinase domain (exon 20, H1047R) of the p110alpha catalytic subunit and determined the effect of PI3K-AKT pathway activation on cellular proliferation, survival and leukemogenic expansion in vitro and in vivo. We demonstrate that the p110alpha point mutations constitutively activated the PI3K-AKT pathway and resulted in factor-independent growth of Ba/F3 cells. Proliferation and survival of the cells were inhibited in a time- and dose-dependent manner using inhibitors of either PI3K or AKT. The mammalian target of rapamycin (mTOR) was demonstrated to be important for mitogenic, but not anti-apoptotic signaling of mutant PIK3CA. In a syngenic mouse model, hematopoietic cells expressing mutant p110alpha induced a leukemia-like disease. FACS analysis demonstrated a median chimerism of 68% in the bone marrow and 35% in peripheral blood of diseased mice, which were characterized by anemia, leukemic infiltration of hematopoietic organs, and 90% mortality within 5 weeks. No substantial differences were observed between E542K, E545A and H1047R. Mice, which were transplanted with an activating c-KIT point mutation (D814V), showed a significantly reduced survival and 100% died within 9 days. In conclusion our data show, that PIK3CA point mutations, by activating the PI3K-AKT pathway, confer factor-independence to hematopoietic cells in vitro and induce leukemogenic activity in vivo. As mutant c-KIT (D814V) showed enhanced leukemogenic potential, signaling through c-KIT may involve additional downstream pathways which cooperate with activated PI3K in leukemia progression. Our model is useful for the differential investigation of the leukemogenic relevance of the PI3K-AKT pathway and pharmacological inhibitor studies.
Phase II Study of High Dose Lenalidomide as Initial Treatment for Older Acute Myeloid Leukemia Patients: Early Results Show a Significant Reduction of Bone Marrow Blasts after 14 Days of Therapy.
Authors: Fehniger Todd A. , Nelson Alissa , Trinkaus Kathryn , Abboud Camille N. , Cashen Amanda F. , DiPersio John F. , Graubert Timothy A. , Ley Timothy J. , Tomasson Michael H. , Uy Geoffrey L. , Walter Matthew J. , Westervelt Peter , Vij Ravi
Published: October 2019

AML patients over the age of 60 years have a poor prognosis, share disease characteristics with myelodysplastic syndrome (MDS) patients, and warrant novel therapeutic approaches. Lenalidomide has immunomodulatory and anti-neoplastic properties which can induce morphologic and cytogenetic responses in MDS patients, including those with excess blasts. We hypothesized that lenalidomide may be active against AML, and have employed a high dose strategy without dose reductions for hematologic toxicities. Here, we report preliminary results from a phase II study of high dose lenalidomide for front-line treatment of AML ≥ 60 without chromosome 5q deletion or favorable cytogenetics. Treatment included 2 cycles of high dose lenalidomide (50mg/day x 14 days, 30 days of rest, 50mg/day x 21 days), followed by maintenance therapy (10 mg/day) in non-progressing patients. Fifteen patients were enrolled in the first stage between 2/27/07 and 8/3/07. Median age was 71 years (range 60–86 years); ECOG performance status was 0 (4/15, 27%), 1 (10/15, 67%), or 2 (1/15, 7%); 11/15 (73%) patients were male; and 5/15 (33%) patients had prior MDS. Cytogenetics were normal (n=9), loss of chromosome 7 (n=2), loss of chromosome 20 (n=1), trisomy 13 (n=1), or complex (n=2). Overall, the treatment regimen was well tolerated. In the first stage of the study, 12/15 patients are evaluable for day 15 bone marrow and peripheral blood blast changes following the initial high dose lenalidomide cycle (50 mg/day x 14 days). Pre-therapy WBC counts (mean ± SEM) were 13,825 ± 4,447/uL (range 1,100–45,300/uL) and day 15 WBC counts were 4,742 ± 2,136/uL (range 300–24,400/uL). Day 15 bone marrow myeloblast percentages were significantly reduced in 9/12 patients (mean ± SEM decrease of 53 ± 10%, P=0.01, range 18–100%). In addition, the bone marrow blast index (% cellularity x fraction of blasts) decreased significantly after 14 days of high dose lenalidomide (mean ± SEM reduction of 66 ± 11%, P=0.02). Moreover, 5/8 patients with circulating blasts at diagnosis showed clearance of their peripheral blasts at day 15. These findings suggest that lenalidomide is an active agent against acute myeloid leukemia. Results on AML blast changes, response by international working group criteria, and toxicities in this patient cohort with ≥4 months of follow-up will be updated and presented.
A New Murine Model of Immune Thrombocytopenia: Evidence of Both Antibody- and CD8+ T Cell-Mediated Platelet Destruction.
Authors: Chow Leola , Ni Heyu , Kim Michael , Speck Edwin R. , Aslam Rukhsana , Chen Pingguo , Freedman John , Semple John W.
Published: October 2019

Immune thrombocytopenic purpura (ITP) is an autoimmune-mediated bleeding disorder in which platelets are opsonized by autoantibodies and prematurely destroyed by Fc receptor (R)-mediated phagocytosis in the reticuloendothelial system (RES). Although the pathogenesis of the disorder has been considered to be primarily antibody mediated, recent reports have indicated that cell-mediated (e.g. CD8+ T cell) mechanisms can also lead to platelet destruction at least in those patients with ITP who have no detectible antibodies. To study these mechanisms, we developed a murine model of immune thrombocytopenia by transferring spleen cells (SC) from GPIIIa-knockout (KO) mice immunized against wild type (WT; GPIIIa-positive) platelets into syngeneic severe combined immunodeficient (SCID) mice. Results show that compared with naive SC, transfer of as few as 5x104 immune GPIIIa-KO SC caused significant thrombocytopenia and a bleeding diathesis and death in 60 percent of recipient SCID mice within 10 days post-transfer. Bleeding occurred primarily in the gut, lungs, subcutaneous tissues and brain. When the immune GPIIIa-KO SC were first depleted of CD4+T cells and transferred, thrombocytopenia or bleeding mortality occurred. In contrast, CD8+ T cell depletion of the GPIIIa-KO SC before transfer did not affect their ability to cause thombocytopenia nor bleeding. Interestingly, depletion of CD19+ B cells before transfer did not affect the ability of the SC to induce thrombocytopenia but prevented all bleeding. These results suggest that both antibody-mediated and CD8+ T cell-mediated platelet destruction occur in this animal model of immune thrombocytopenia, however, it appears that only antibody-mediated thrombocytopenia is associated with bleeding mortality. Because the immune thrombocytopenia is against a platelet-specific antigen e.g. GPIIIa (like in ITP), it will not only allow for the study of disease pathogenesis but may also be important for testing new immunospecific therapeutics to increase platelet counts in patients with ITP.
Mantle Cell Lymphoma Can Be Cured by Intensive Immunochemotherapy with In-Vivo Purged Stem-Cell Support; Final Report of the Nordic Lymphoma Group MCL2 Study.
Authors: Geisler Christian H. , Elonen Erkki , Kolstad Arne , Laurell Anna , Andersen Niels , Pedersen Lone B. , Boesen Anne Marie , Eriksson Mikael , Jerkeman Mats , Kimby Eva , Kuittinen Outi , Lauritzen Grethe F. , Nilsson-Ehle Herman , Nordström Marie , Ralfkiaer Elisabeth , Åkerman Måns , Ehinger Mats , Sundström Christer , Langholm Ruth , Delabie Jan , Karjalainen-Lindsberg Marja-Liisa , Brown Peter
Published: October 2019

Mantle cell lymphoma (MCL) is considered incurable, with a median survival of 4 years. Intensive immunochemotherapy and autologous stem-cell (ASC) support has appeared promising in small patient cohorts, but has not been tested in large, consecutive series. Here we report the final results of the 2nd Nordic MCL (MCL2) trial after a median of 3 years follow-up from study entry.

Methods: This unrandomized phase-II trial included 159 untreated patients younger than 66 years, 84% stage IV, 128 with classical, 31 with blastoid/pleomorphic cytology. Following 6 cycles of intensive induction immunochemotherapy with alternating cycles of rituximab (R) + maxi-CHOP and R+ high-dose AraC, responders received BEAM/BEAC with in-vivo purged (R) ASC support.

Results: 153 patients (96%) responded to induction therapy with CR in 55% and PR in 41%. The 5-year event-free (EFS) and overall survival (OAS) are 63% and 74% respectively on intention-to-treat, and the 144 (91%) responders who completed treatment had 72% 5-year response duration, with plateaus emerging in all three curves at these levels.

Figure Figure

There were 6 treatment-related deaths (3,8%). Of 77 patients with available primers, 90% had become PCR-negative two months posttransplant; those who remained PCR-negative more than 1 year posttransplant had a significantly longer clinical response duration than patients who did not (P<0.0001). Of 42 stem-cell products assessed 88% were PCR-negative as compared to only 12% in the MCL1 study (P<0.001). In a multivariate analysis including age, sex, international prognostic index (IPI), cytological variants and Ki-67 proliferation index, only IPI and Ki-67 were independent predictors of EFS and response duration respectively, and only IPI and cytological variant of OAS. Compared to the 41 patients of 1st Nordic MCL study who received 4 cycles of maxi-CHOP without rituximab before BEAM or BEAC + ASCT (1), the OAS, EFS and response duration were highly significantly increased.

Conclusion: The demonstration of long-term event-free survival in a large, consecutive prospective series now for the first time indicates that intensive immunochemotherapy including AraC and Rituximab with in-vivo purged stem-cell support may cure mantle cell lymphoma.
Cardiac Toxicity Associated with the Anti-VEGF Monoclonal Antibody Bevacizumab (Avastin) in Combination with CHOP (A-CHOP) Chemotherapy for Peripheral T Cell Lymphoma (PTCL): The ECOG 2404 Trial.
Authors: Advani Ranjana H , Hong Fangxin , Ganjoo Kristen N. , Manola Judith B. , Swinnen Lode J. , Habermann Thomas M , Horning Sandra J
Published: October 2019

Abstract 1671

Poster Board I-697

Background

Bevacizumab (Avastin), a humanized monoclonal antibody directed against vascular endothelial growth factor (VEGF-A), is approved in combination with chemotherapy in lung, colorectal, breast and brain cancers. VEGF-A expression is reported in all lymphoma subtypes, with strongest expression in PTCL. Given the relatively poor prognosis of PTCL with standard therapy, we combined bevacizumab (A) with CHOP in the ECOG 2404 phase II trial.

Methods

Untreated PTCL pts with a normal baseline cardiac ejection fraction received bevacizumab 15 mg/ kg IV and CHOP chemotherapy on day 1 for 6-8 cycles. Patients (pts) with CR/PR/SD received maintenance therapy with bevacizumab 15 mg/ kg IV q 3 weeks for 4 cycles. Due to the reported 3.7% incidence of cardiac toxicity associated with bevacizumab in metastatic breast cancer pt previously exposed to anthracyclines (Avastin package insert), cardiac function was monitored prospectively with ejection fraction (EF) determination at the completion of A-CHOP prior to the start of bevacizumab maintenance.

Results

Among 33 pts who came off study, toxicity data are available for all and coded using CTCAE Version 3.0. 22 received at least 4 cycles of A-CHOP (median treatment duration 3.6 months (< 1-10.6 months). One or more designated cardiac toxicities were recorded for 5 of these 22 pts. Congestive heart failure (CHF), defined as grade 2-4 left ventricular dysfunction was recorded in 4 pt (18%, 90% CI 5.2%-40.3%): grade 4 (n=1), grade 3 (n=3). In 3 pts clinical CHF occurred after A-CHOP x 6 and in one during bevacizumab maintenance after A-CHOP x 8. The median time to development of a cardiac adverse event was 5 months (3.5 – 8.8 months). One pt required a ventricular assist device in addition to medical management. In 3 of the 4 pts, ongoing medical management for CHF has been required. Other cardiac toxicities included ventricular arrhthymia (one grade 4, one grade 2), and one grade 2 cardiac troponin elevation. In addition, one patient experienced sudden death of uncertain cause. Three pts developed thromboses or embolic toxcities (two grade 4 and one grade 3).

Conclusions

We observed four cases of clinical CHF early after cumulative doxorubicin doses of 300-400 mg/m2 in our study, The incidence of CHF among study patients who received 4 or more A-CHOP cycles is concerning and suggests that bevacizumab may potentiate the adverse cardiac effects of anthracyclines, resulting in clinically significant toxicity. Careful monitoring of cardiac function is warranted in ongoing trials combining bevacizumab and doxorubicin in lymphomas and solid tumors to assess long-term safety and better understand underlying mechanisms and risks.

Disclosures

No relevant conflicts of interest to declare.
CD40 Ligand Determines Whether Interleukin 21 Induces Differentiation of Human B Cells Into Plasma Cells or Into Granzyme B-Secreting Cytotoxic Cells.
Authors: Jahrsdoerfer Bernd , Hagn Magdalena , Sontheimer Kai , Schwesinger Elisabeth , Ebel Verena , Maier Julia , Beyer Thamara , Fabricius Dorit , Lunov Oleg , Tron Kyrylo , Syrovets Tatjana , Barth Thomas FE , Nienhaus G. Ulrich , Simmet Thomas
Published: October 2019

Abstract 2675

Poster Board II-651

Interleukin 21 (IL-21) is a novel and highly promising cytokine for the treatment of neoplastic and infectious diseases. Recently, IL-21 has been identified as inducer of plasma cell differentiation. Here we show that CD40 ligand is critically involved in this process and that, in its absence, human B cells differentiate into granzyme B (GzmB)-secreting cytotoxic cells rather than plasma cells. GzmB expression and secretion by human B cells was demonstrated by FACS analysis, ELISpot, ELISA, Sensizyme, Western immunoblotting, RT-PCR, and spinning disk confocal microscopy. GzmB secretion requires the presence of IL-21 and B cell receptor engagement, and depends on phosphorylation of JAK1/3 and STAT3. CD40 ligation effectively suppresses GzmB secretion by B cells, suggesting GrB-secreting B cells play a role in the early phase of inflammatory processes, before CD40 ligand-expressing T cells are present. Of note, ex-vivo re-stimulation of B cells from recently vaccinated individuals with inactivated viruses also induces GzmB expression. GzmB is enzymatically active and GzmB-secreting B cells induce apoptosis in various tumor cell lines, a process we were able to visualize by using spinning disk confocal microscopy. Our data reveal an as yet unrecognized role of IL-21-activated B cells, which involves GzmB secretion and cellular cytotoxicity. Our findings may have implications for the understanding of tumor immunosurveillance and early anti-viral immune responses, and may open novel approaches for the immunotherapy of neoplastic and viral diseases.

Disclosures:

No relevant conflicts of interest to declare.
Asymptomatic Familial Factor XI Deficiency.
Authors: Herrada Juan , Aloba Maria Chona , Adcock Dorothy M , Gupta Anuradha , Noble Luis S. , Landeros Mark M.
Published: October 2019

Abstract 4449

Background

Factor XI deficiency (known as Rosenthal syndrome or hemophilia C) is an autosomal disorder affecting both sexes which results in a bleeding disorder of variable severity. This condition is very uncommon among the non-Jewish population and consists mostly of sporadic cases, although an occasional familial cluster has been described (Bolton-Maggs et al. J Thromb Haemost 2004 Jun;2(6):918-24). We present two asymptomatic siblings with coagulation Factor XI deficiency.

Case Report

In February 2004 a healthy 33 year-old non-Jewish white female without personal or family medical history presented with an elevated partial thromboplastin time (PTT) incidentally found during a routine pre-operative evaluation. Her prothrombin time (PT) was normal. Her PTT was 45 seconds (reference interval 23-37 seconds), and normalized with 1:1 mixing studies of patient and normal plasma. Preliminary laboratory evaluation included negative tests for the presence of a Lupus Anticoagulant, a normal serum albumin level, and a normal urinalysis. Additional laboratory testing of coagulation and included FXI and XII activities, measured by means of a one stage APTT-based clotting assay using APTT reagent (Automated APTT, Trinity Biotech, Bray Ireland), congenitally depleted deficient plasma (HRF, Inc, Raleigh NC) using the MDA II Trinity Biotech, Bray Ireland). FXI was 46% (reference range 60-150) and FXII was 42% (reference range 50-150). In March 2004, a repeated the laboratory evaluation showed a FXI of 41% and a FXII of 46%. In April 2006, FXI was 49%, and FXII was 40%. Is to be noted that the patient never had abnormal bleeding episodes despite abdominal and breast surgical procedures. In 2009, her only brother (aged 29 and otherwise healthy) underwent hematological evaluation that showed normal levels of PT, PTT, platelet count, fibrinogen, and factor XII (88% activity). His factor XI was 51% (decreased activity).

Conclusion

Although modern laboratory methods are able to identify unusual coagulation defects, their clinical significance requires additional investigation.

Disclosures:

No relevant conflicts of interest to declare.
Anthracycline Treatment of the Monocytic Leukemia Cell Line THP-1 Increases Phosphatidylserine Exposure and Tissue Factor Activity.
Authors: Boles Jeremiah , Glover Sam , Owens A. Phillip , Hollingsworth Rachel , Kasthuri Raj S. , Key Nigel S. , Mackman Nigel
Published: October 2019

Abstract 1136

Introduction:

Cancer associated thrombosis is a well-recognized phenomenon that results in considerable patient morbidity and mortality. While malignancy alone conveys an increased risk for thrombosis, treatment with cytotoxic chemotherapeutics further elevates this risk. However, the pathogenic mechanisms underlying this process remain poorly defined. We hypothesized that treatment of tumor cells with cytotoxic chemotherapeutic drugs leads to phosphatidylserine (PS) exposure on the cell surface, as well as increased cellular tissue factor (TF) activity and release of TF positive microparticles (MPs).

Aim:

The aim of this study was to determine the effect of anthracyclines on TF activity and MP release from the human monocytic leukemia cell line THP-1.

Materials and Methods:

THP-1 cells were treated with different concentrations of doxorubicin (0-10μg/ml) or daunorubicin (0-1μg/ml) for 0–24 hours. These concentrations are similar to those found in plasma of cancer patients. The intact cells were analyzed for TF activity using a two stage chromogenic assay. TF antigen was evaluated by ELISA and TF mRNA by realtime PCR. In addition, TF activity of MPs released into the culture supernatant was measured using a two stage chromogenic assay. PS exposure on cells and MPs were analyzed by flow cytometry and PS exposure on MP was also evaluated in an annexin V capture assay. Cell viability was assayed by a dye exclusion assay as well as by flow cytometry.

Results:

We found that doxorubicin and daunorubicin increased PS exposure on the cell surface. Treatment with anthracyclines increased both cellular TF and MP TF activity in a concentration-dependent manner that also resulted in concentration dependent increase in apoptosis. Furthermore, this increased TF dependent activity was abolished with the addition of annexin V or lactadherin supporting a role of PS exposure in this elevated activity.

Conclusion:

These results support our hypothesis that treatment of THP-1 cells with chemotherapeutic agents induces apoptosis and increases cellular TF activity. This increased activity appears primarily mediated by an increase in cell surface expression of PS. Additionally, the anthracyclines increase the release of TF positive MPs from the THP-1 cells. We propose that this increase in cellular TF activity in tumor cells and increased numbers of TF positive MPs may contribute to thrombosis in cancer patients treated with chemotherapeutic drugs.

Disclosures:

No relevant conflicts of interest to declare.
Relationship of Soluble Adhesion Receptors to Red Cell Apoptosis In SCD.
Authors: Mutucumarana Charmaine P. , Chandler Catherine , Telen Marilyn J. , Delahunty Martha
Published: October 2019

Abstract 1654

The interaction of sickle red cells (SS RBCs) with endothelium can cause endothelial injury, leading to increased expression of soluble endothelial cell adhesion molecules and plasma markers of endothelial activation and dysfunction. Among such plasma markers, sVCAM-1 levels are increased in SCD and rise further during vaso-occlusive crisis (Duits et al. 1993), and several groups have shown that sVCAM-1 levels are positively related to LDH levels, a marker of hemolysis. Perfusion of endothelial cells with SS RBCs has been shown to increase the expression of the cellular adhesion molecules ICAM-1 and VCAM-1 by endothelial cells (Shiu et al. 2000). Furthermore, sVCAM-1, sICAM-1, and sE-selectin have been independently associated with risk of mortality (Kato et al. 2005). We were interested in determining whether identifiable characteristics of SS RBCs themselves might be associated with the degree to which soluble adhesion molecules are expressed in SCD patient plasma. Therefore, we measured sVCAM-1, sICAM-1, sCD40 ligand (sCD40L) and sP-selectin in SCD patient plasma by ELISA (n=46) and looked at the relationship of these biomarkers of disease activity to SS RBC characteristics. We specifically examined phosphatidylserine exposure (as measured by annexin V binding analyzed by flow cytometry) and the degree of SS RBC adhesion to laminin and endothelial cells, measured in vitro in a graduated height flow chamber. We found that the degree of red cell damage measured by annexin V binding correlated with the level of soluble adhesion receptors sVCAM-1 (p=0.036) and sICAM-1 (p=0.026). However, annexin V binding did not correlate with sCD40L or sP-selectin expression in plasma, and neither annexin V binding nor levels of soluble adhesion molecules correlated with SS RBC adhesion to laminin or endothelial cells in vitro. In addition, the concentrations of sVCAM-1 and sICAM-1 in plasma were strongly correlated with each other (p<0.0001), suggesting that both of these adhesion molecules may be released from activated endothelial cells in SCD patients as a result of the same mechanism involving contact with phosphatidylserine-exposing SS RBCs. These data support the hypothesis that phosphatidylserine exposure on SS RBCs is a critical factor in SS RBC interaction with the endothelium (Setty et al., 2002), including the release of soluble cellular adhesion markers that indicate endothelial activation and/or injury.

Disclosures:

Telen: GlycoMimetics, Inc.: Consultancy.
12-Month Follow-up for Patients with Sickle Cell Disease in an Ongoing 3-Year, Prospective, Non-Interventional Registry Trial
Authors: Heeney Matthew M. , Mueller Brigitta U. , Baltz Brad , Adams-Graves Patricia , Yang Elizabeth , Paley Carole , Esposito Jason , Vichinsky Elliott P.
Published: October 2019

Abstract 1010

Introduction:

Sickle cell disease (SCD) is a clinically heterogeneous disease. Patients (pts) may present with severe symptoms in childhood, resulting in early morbidity and mortality, or may be asymptomatic until adulthood. Therapeutic goals include the prevention and treatment of complications across the spectrum of disease. We describe baseline and 12-mo assessments and therapies for pts in an ongoing registry documenting treatment patterns, natural history, and outcomes.

Methods:

Pts ≥2 years old with HbSS, HbS/β-thalassemia, or HbSC were enrolled from 57 US centers and assessed every 6 mos for up to 3 years. Differences between pediatric (age <18) and adult (age ≥18) pts at 12 mos follow-up are reported. (ClinicalTrials.gov identifier, NCT01220115.)

Results:

In all, 498 pts completed the baseline visit. Most pts had HbSS disease (age <18, 76.3%; age ≥18, 70.2%). Adult pts had worse performance status, and more frequent avascular necrosis, gallbladder disease, leg ulcers, and pulmonary hypertension at baseline (Table 1). They also had more transfusions, higher serum ferritin, more chelation exposure, more hospitalizations for pain, and potentially more strokes (Table 2). Pediatric pts had higher frequencies of asthma, abnormal transcranial Doppler (TCD) readings, dactylitis, splenic sequestration, acute chest syndrome, infections, hospitalizations at baseline, and more prevalent hydroxyurea use on study. Absenteeism from school (age <18, 1 to 10 d missed prior to study, 50.8%; 1 to 10 d missed on study, 65.8%) and work (age ≥18, 1 to 10 d missed prior to study, 44.4%; 1 to 10 d missed on study, 45.1%) was frequent.

Conclusions:

This study showed differences in baseline characteristics and treatment patterns between adult and pediatric pts with SCD. Ongoing follow-up will provide further information on disease patterns, treatment practices, and outcomes, thus increasing our understanding of appropriate SCD management.

Disclosures:

Heeney: Novartis: Consultancy, Research Funding; Eli Lilly: Research Funding. Off Label Use: Hydroxurea is indicated to reduce the frequency of painful crises and to reduce the need for blood transfusions in adult patients with sickle cell anemia. It is not approved for use in children. Mueller:Novartis: Research Funding. Adams-Graves:Novartis: Consultancy, Speakers Bureau. Paley:Novartis: Employment. Esposito:Novartis: Employment. Vichinsky:Novartis: Consultancy, Research Funding; ApoPharma: Consultancy, Research Funding; ARUP Research Laboratory: Research Funding.
Novel 2-Phenyl-1-Pyridin-2yl-Ethanone (PpY) Based Iron Chelators Increase Expression of IkBα and Heme Oxygenase-1 and Inhibit HIV-1
Authors: Kumari Namita , Xu Min , Kovlaskyy Dmytro , Dhawan Subhash , Nekhai Sergei
Published: October 2019

Abstract 1052

HIV-1 transcription is activated by HIV-1 Tat protein, which recruits CDK9/cyclin T1 and other host transcriptional co-activators to the HIV-1 promoter. Tat itself is phosphorylated by cell cycle kinase 2 (CDK2) and inhibition of CDK2 by small interfering RNA or iron chelators inhibits HIV-1 transcription. HIV-1 transcription is also activated by NF-kB that binds to HIV-1 LTR independent to Tat but can also be recruited Tat-dependently by CDK9/cyclin T1. Recently, induction of heme oxygenase-1 (HO-1) by hemin was shown to inhibit HIV-1 in vitro and in vivo. Here, we analyzed the effect of novel phenyl-1-pyridin-2yl-ethanone (PPY) based iron chelators, PPYeT and PPYaT, on HIV-1. Both chelators efficiently inhibited one round of HIV-1 replication in T cells at low nanomolar IC50s without exhibiting cytotoxicity at 24 hrs incubation. The iron chelators efficiently bound intracellular labile iron as it was determined in calcein binding assays. Because we previously showed that iron chelators inhibited the activity of CDK9, we analyzed expression of several cellular genes dependent on CDK9. Unexpectedly, chelators were found to induce the expression of IkBα, an inhibitor of NF-kB (Fig1). Treatment with the iron chelators retained NF-kB in cytoplasm of the treated cells suggesting reduction in NF-kB in nucleus (Fig2). The chelators were also shown to induce HO-1 expression in cultured monocytes, likely to do a decrease of intracellular iron pool. This effect of iron chelators mimicked the effect of hemin treatment which also induced HO-1 and inhibited HIV-1 infection in our experimental conditions. Low nanomolar IC50s for the PPY-based iron chelators and lack of toxicity suggest their potential usefulness as future anti-retroviral therapeutics. Further studies are needed to investigate additional targets for iron chelators in HIV-1 life cycle that may include reverse transcription and capsid assembly. Therefore iron chelators need to be carefully assessed not only to understand the mechanism but also as a therapeutic strategy.

Acknowledgments.

This work was supported NIH Research Grants SC1GM082325, R25 HL003679, 2G12RR003048, 8G12MD007597, K25GM097501 and 1P30HL107253.

Disclosures:

No relevant conflicts of interest to declare.
Genome-Wide Association Study of Platelet Function in African Americans
Authors: Qayyum Rehan , Becker Lewis , Becker Diane , Yanek Lisa , Leal Suzanne M , Mathias Rasika , Bray Paul F.
Published: October 2019

Abstract 1068

Background:

We have previously shown that platelet aggregation, an essential component of pathogenesis of acute coronary syndromes, has higher heritability in African Americans than Caucasians. However, a genome-wide association study of native platelet function in African Americans has not been reported.

Methods:

Platelet-rich plasma (PRP) was isolated from blood samples obtained from the discovery (GeneSTAR, N=835) and replication (Platelet Genes and Physiology Study, N=119) cohorts. Optical aggregation was used to measure maximal aggregation in PRP after samples were stimulated with arachidonic acid, collagen, ADP, or epinephrine. Genotyping was conducted with Illumina 1M arrays. For each cohort, age- and sex-adjusted linear mixed models were used to test for association between each SNP and each phenotype under an additive model.

Results:

Of the 117 SNPs that were significant (P< 5 × 10−8) in the discovery sample, 50 SNPs in 4 regions were also significant in the replication sample (Table). Among the replicated SNPs, a previously reported SNP in European Americans, rs12041331, in the PEAR1 gene was associated with ADP- and epinephrine-mediated aggregation. 47 SNPs in the 17q21.31 region, in one LD block, were associated with collagen-mediated aggregation.

Conclusions:

In this first GWAS of native agonist-mediated platelet aggregation in African Americans, we have discovered, and replicated in an independent sample, 4 regions that are associated with platelet aggregation. Several genes in the identified regions are expressed in platelets and further study of these genes may provide novel insights in platelet biology.

Disclosures:

No relevant conflicts of interest to declare.
Direct Observation of Von Willebrand Factor Elongation and Fiber Formation On Collagen During Acute Whole Blood Exposure to Pathological Flow
Authors: Colace Thomas Vincent , Diamond Scott L.
Published: October 2019

Abstract 1070

Under conditions of pathological shear rate, von Willebrand Factor (vWF) undergoes conformational changes and self aggregation. We sought to visualize this phenomenon using a novel microfluidic model of stenosis and understand its role in thrombus formation in elevated shear rate environments. In severe stenosis, vWF experiences millisecond exposures to pathological wall shear rates (gw). Distinct from shear experiments that last many seconds, we deployed microfluidic devices for single-pass perfusion of whole blood or platelet free plasma (PFP) over fibrillar type 1 collagen (< 50 msec transit time) at pathological gw or spatial wall shear rate gradient (grad gw). Using fluorescent anti-vWF, long thick vWF fibers (>20 mm) bound to collagen were visualized at constant gw > 30,000 s−1 during perfusion of PFP, a process enhanced by EDTA. Rapid acceleration or deceleration of EDTA-PFP at grad gw = ± 5.5 × 105 to 4.3 × 107 s−1/cm did not promote vWF deposition when gw < 30,000. At 19,400 s−1, EDTA-blood perfusion resulted in rolling vWF-platelet nets, while blood perfusion (normal Ca2+) generated large vWF/platelet deposits that repeatedly embolized and were blocked by anti-GP1b or the aIIbβ3 inhibitor GR144053 and did not require shear gradients. Blood perfusion at venous shear rate (200 s−1) produced a stable platelet deposit that was a substrate for massive but unstable vWF-platelet aggregates when flow was increased to 7800 s−1. Supported by collagen and enhanced by platelet GP1b and aIIbβ3, vWF fiber formation occurred during acute exposures to pathological gw but did not require wall shear rate gradients.

Figure 1 A, Platelet free citrated-plasma was treated with 1 μg/mL fluorescently labeled anti-vWF and 5 mM EDTA. The plasma samples were perfused over a collagen type 1 surface at local wall shear rates of 30,000, 62,400 and 125,000 s−1 from left to right. Long fibers of vWF (>20 μm) appeared at shear rates above ∼30,000 s−1, with more fibers appearing at higher shear rates. The bar indicates 15 μm. Figure 1. A, Platelet free citrated-plasma was treated with 1 μg/mL fluorescently labeled anti-vWF and 5 mM EDTA. The plasma samples were perfused over a collagen type 1 surface at local wall shear rates of 30,000, 62,400 and 125,000 s−1 from left to right. Long fibers of vWF (>20 μm) appeared at shear rates above ∼30,000 s−1, with more fibers appearing at higher shear rates. The bar indicates 15 μm.

Disclosures:

No relevant conflicts of interest to declare.