bluuebunny/crossref_metadata_2025 huggingface dataset:
Prevalence of Thrombophilia in Women with Spontaneous Pregnancy Loss.
Authors: De Sancho Maria Teresa , Richardson Lamar , Rand Jacob H.
Published: October 2019

We retrospectively analyzed the records of 145 female patients ages 18 to 50 who were referred to the Thrombosis and Hemostasis Section of the Hematology-Oncology Division at Mt. Sinai Hospital from January 2000 to June 2004 for evaluation of thrombophilia as the etiology of pregnancy loss (PL).

Recurrent Pregnancy Loss (RPL) was defined as 2 or more PL in the first trimester of gestation. Women with one or more PL in the second and/or third trimester of gestation or stillbirth, or a combination of above were also analyzed.

The following thrombophilic conditions were evaluated: factor V Leiden, prothrombin gene mutation (G20210), antiphospholipid (APL) antibodies and lupus anticoagulant (LAC), homocysteine and functional levels of protein C, S and antithrombin.

Of the 145 patients, 20 were excluded from analysis for the following reasons: isolated 1 first trimester PL (15 patients), no verification of antiphospholipid antibodies (4 patients) and insuficient data (1 patient).

Of the evaluable patients, 75 (60%) had 2 or more first trimester PL, 14 (11.2%) patients had 1 or more second trimester PL, 2 (1.6%) patients had 1 or more third trimester PL and 24 (19.2%) patients had PL in different trimesters.

70 of the 125 patients (56%) were found to have a thrombophilic condition: 25 had antiphospholipid antibodies, 21 had factor V Leiden (20 heterozygous and 1 homozygous), 16 had prothrombin gene mutation (all heterozygous), 5 had protein S deficiency, 2 had homocysteine level, and 1 had protein C deficiency. We did not identify any patient with antithrombin deficiency.

Of the 75 patients with first trimester PL, 36 (48%) were found to have a thrombophilic condition: 18 (50%) had antiphospholipid antibodies, 9 (25) had factor V Leiden (1 homozygous and 8 heterozygous) (17%), 5 (13.8) had heterozygous prothrombin gene mutation (8%), 3 (8.3%) had increased homocysteine level and 3 had a low protein S level (8.3%).

In summary, we found a thrombophilic condition associated with PL in 56% in this patient population. The majority of patients had recurrent first trimester PL. The most prevalent thrombophilic disorder was the presence of antiphospholipid antibodies, followed by factor V Leiden, prothrombin gene mutation and protein S deficiency and the least common was increased homocysteine level. These results suggest that testing for thrombophilia may be warranted in women with recurrent PL.
Pegfilgrastim Alone Successfully Mobilizes Peripheral CD34+ Cells in Chemotherapy Naive Subjects with Solid Tumors: Initial Results of a Phase 1-2 Study.
Authors: Willis Fenella , Pettengell Ruth , Woll Penella J. , Desborough Claire , Buchanan Karen
Published: October 2019

BACKGROUND: Classic strategies for peripheral blood progenitor cell (PBPC) mobilization include daily administration of a growth factor, such as filgrastim, alone or with marrow suppressive chemotherapy. A single injection of pegfilgrastim has been shown to be comparable to daily injections of filgrastim in the treatment of chemotherapy-induced neutropenia. One of the objectives of this phase 1–2 study was to provide dose-finding information regarding the efficacy and kinetics of cytokine alone PBPC mobilization with pegfilgrastim.

METHODS: Chemotherapy naïve subjects with a variety of solid tumors were randomized to receive a single administration of 6, 12 or 18 mg of pegfilgrastim on day 1 or daily administration of 10 μg/kg filgrastim from day 1 until day 7. Daily blood samples for peripheral CD34+ analysis were collected from days 1 to 7 and then on days 9 and 12. No chemotherapy was administered during this cycle. For each subject, peak CD34+ cell count was defined as the maximum cell count observed between days 3 to 7, inclusive.

RESULTS: Of 61 subjects randomised into the study, all received study drug (15 subjects each in the pegfilgrastim 12mg, 18mg and filgrastim groups and 16 subjects in the pegfilgrastim 6mg group). The treatment groups were balanced in terms of demographics and baseline characteristics. The most common tumor types were non-small cell lung cancer (n= 23 [38%]) and ovarian (n= 19 [31%]). The mean peak CD34+ cell count was similar in the filgrastim and pegfilgrastim 6mg groups (4.51 and 4.24 x 104/mL respectively) whereas the pegfilgrastim 12mg and 18mg groups had higher mean peaks (8.18 and 9.96 x 104/mL respectively). Pegfilgrastim 12mg and 18mg mobilized significantly more peripheral CD34+ cells than filgrastim (p= 0.025 and 0.003 respectively). The subject incidence of SAEs was low and comparable between groups.

Figure Figure

CONCLUSION: In chemotherapy naïve subjects with solid tumor, single day administration of pegfilgrastim is at least as efficacious as repeated daily injections of filgrastim (10 μg/kg) in its ability to successfully mobilize peripheral CD34+ cells. A dose response relationship of CD34+ mobilization is observed. Pegfilgrastim 12mg and 18mg mobilize higher numbers of CD34+ cells than daily administrations of filgrastim. Pegfilgrastim at 6, 12 and 18mg is generally safe and well tolerated in this setting.
Evidence for Safe Exclusion of Deep Vein Thrombosis (DVT) by a Rapid ELISA D-Dimeer Test at Cut-Off Levels of 500 and 1000 Ng/Ml in 1046 Consecutive Outpatients with Suspected DVT.
Authors: Michels Jan Jacques , Hermssen Jan , Trienekens Paul H.
Published: October 2019

Introduction.A normal compression ultrasonography (CUS) safely excludes proximal deep vein thrombosis (DVT) with a negative predictive value of 97% indicating the need to repeat CUS testing within one week. In 3 studies, the rapid ELISA D-dimer assay at a cut-off of 500 ng/ml did have a sensitivity of 100% for the exclusion of venographically documented distal and proximal DVT irrespective of clinical score. To test this hypothesis we performed a large prospective study in outpatients with suspected DVT.

Methods. CUS and a rapid ELISA D-Dimer test (VIDAS, BioMérieux L’Etoile, France) were performed in patients with suspected DVT. A negative CUS with a D-Dimer result of <500 ng/ml exclude DVT, and with a D-Dimer result of >500 ng/ml was followed by a second CUS within one week.

Results. The prevalence of DVT 1046 consecutive out patients with suspected DVT was 23,4%. The first CUS was positive in 228 with a rapid ELISA D-Dimer of >500 ng/ml in 227 and of <500 ng/ml in one case, indicating a sensitivity of 99,6% irrespective of clinical score. The first CUS was negative in 818. The rapid ELISA D-dimer test Was <500 ng/ml in 297 of which 296 had a negative first CUS indicating a negative predictive value of 99.7% at a specificity of 37% irrespective of the clinicl score. The negative predictive value of a negative CUS plus a rapid ELISA D-Dimer result of less than 1000 ng/ml is 99.5% at a specificity of 67,9% irrespective of clinical score. The prevalence of DVT in patients with negative first CUS and a ELISA D-Dimer of >1000 ng/ml was 5.6% as documented by CUS repeat within on week.

Conclusion.A normal rapid ELISA D-dimer test, <500 ng/ml, in outpatients with suspected DVT safely excludes DVT irrespective of clinical score. After a negative rapid ELISA result (<500 ng/ml), CUS is still indicated for safety reasons in patients with suspected DVT and persistent symptoms in search for an alternative diagnosis, or for a rare case of DVT. A negative CUS plus and ELISA D-Dimer result of <1000 ng/ml safely exclude DVT without the need to repeat CUS in 2/3 of patients with a negative first CUS.
Common Polymorphisms of the Interleukin-6 and Chitotriosidase Genes Are Associated with the Risk for Infection with Gram-Negative Bacteria in Children Undergoing Therapy for Acute Myeloid Leukemia.
Authors: Lehrnbecher Thomas , Bernig Toralf , Hanisch Mitra , Koehl Ulrike , Reinhardt Dirk , Creutzig Ursula , Chanock Stephen J. , Klingebiel Thomas , Schwabe Dirk
Published: October 2019

Purpose: Infectious complications remain a substantial cause of morbidity and mortality in patients with acute myeloid leukemia (AML). Candidate gene studies have identified common polymorphisms in genes of molecules of innate immunity as risk factors for susceptibility and/or outcome to a spectrum of pathogens in various populations. This study was performed to investigate whether common polymorphism in genes of innate immunity might influence the risk for serious infection during therapy for pediatric AML.

Patients and Methods: Genotype analysis was performed in 168 children treated on clinical trial AML-BFM 93. Infectious events were categorized as microbiologically or clinically documented infections and as fever without an identifiable source. Candidate gene polymorphisms included tumor necrosis factor-alpha, interleukin 6 and 8 (IL6 and IL8), myeloperoxidase, chitotriosidase (CHIT), the low affinity Fc receptor 2A, and the toll like-receptors 2 and 4.

Results: The 168 children treated according to clinical trial AML-BFM 93 experienced a total of 508 infectious episodes. Only three patients escaped significant infectious complication. One-hundred-and-ten patients had a least one microbiologically documented infection. Gram-positive bacteria were isolated in the bloodstream of 84 children, whereas Gram-negative bacteria were recovered from the bloodstream of 24 patients. There was an association between Gram-negative bacterial infection and each of the IL6 and CHIT genetic variants. The risk of bacteremia with Gram-negative organisms was significantly higher in patients with the G allele in the IL6 promoter at 174bp [22/123 (17.9%) vs 0/25 (0%); P=.022 (OR 11.31, 95% CI 1.41–90.6)] and in patients with the H allele (24-bp duplication in exon 10) of CHIT [13/56 (23.25) vs 11/112 (9.8%); P=.020 (OR 2.78; 95% CI 1.18–6.54)]. No significant association of the other variant genotypes with any of the clinical endpoints analyzed was observed.

Conclusion: The data suggest that variant alleles of both IL6 and CHIT influence susceptibility to infection with Gram-negative bacteria in children undergoing therapy for AML. Further studies are warranted to confirm our findings and to investigate underlying mechanisms.
Comparison of Diagnostic and Relapse Flow Cytometry Phenotypes in Childhood ALL: Implications for Residual Disease (MRD) Detection.
Authors: Borowitz Michael J. , Pullen D. Jeanette , Winick Naomi , Martin P. L. , Bowman W. Paul , Camitta Bruce
Published: October 2019

Flow cytometric analysis of MRD depends on detecting phenotypically abnormal populations. Phenotypic shifts are known to occur but there is little information to indicate how these shifts affect detection of MRD. We compared diagnostic and relapse bone marrow specimens in 36 children with precursor B-ALL studied with the two-tube panel CD19-APC/CD45-PerCP/CD10-PE/CD20-FITC and CD19/45/9/34 (Leukemia 13:558,1999). Small phenotypic shifts of either intensity or coefficient of variation of the distribution of at least one marker were seen in 34 cases, while 7 cases had shifts of 4 or more markers. Phenotypic shifts were complex and could not be explained simply by a change in maturation stage. CD19 was the most stable marker, changing in only 8 cases, CD45 changed in 10; CD10 in 13; CD9 in 14; CD20 in 16 and CD34 in 18. CD10 and CD20 generally lost intensity while CD45 mostly increased. CD34 changed from positive to negative (5 cases) or relapsed as the CD34-negative component when the diagnostic sample was partially positive (9 cases). In 26 cases there was a sufficient change in phenotype from diagnosis to relapse to determine if day 28 MRD more closely resembled the initial diagnostic or the subsequent relapse sample. In 13/19 MRD+ cases the day 28 phenotype resembled the diagnostic specimen and in 6 it resembled relapse. Surprisingly, time to relapse was similar in both groups. In 6 of the 7 MRD negative cases, retrospective review of the data did not identify an abnormal population that resembled either diagnosis or relapse. In the remaining case, in which both CD34 and CD10 were lost from diagnosis to relapse, it is possible that a MRD population resembling relapse might have been missed. We conclude that phenotypic shifts between diagnostic and relapse specimens in childhood ALL are relatively common, but that in general they do not interfere with the ability to recognize MRD. However, MRD analysis that adheres to rigid gating looking only for the precise abnormal phenotype seen at diagnosis might miss some positive cases.
Cyp33-MLL Interaction Modulates Human HOX Gene Expression and Chromatin Structure.
Authors: Diaz Manuel O. , Koonce Mark A.
Published: October 2019

The MLL gene is involved in translocations and fusion genes associated with human leukemia. We have identified a protein, Cyp33, which interacts with the PHD fingers of MLL, and modulates the expression of HOX genes regulated by MLL. In order to define possible mechanisms for this modulation, we set up an experimental system in which the Cyp33 protein is conditionally expressed from a transgene under the control of a tetracycline inducible promoter. Two different cell lines with such inducible gene were established, based on the human kidney epithelial cells HEK293, and the human lymphoblastic leukemia cell line Jurkat, respectively. In these systems we have shown that induction of Cyp33 expression over endogenous levels, leads to downregulation of the HOXC8 gene, but this downregulation is reversible, the HOXC8 being re-expressed when Cyp33 is de-induced and its levels go back to normal. This indicates that Cyp33 overexpression does not erase an epigenetic mark for HOXC8 gene expression. Concordantly, no change in methylation of histone H3 at lysines 4 or 9 is observed upon Cyp33 overexpression. We also showed that both MLL and Cyp33 bind to the promoter of HOXC8, and that histones H3 and H4 at this promoter are de-acetylated upon Cyp33 overexpression. This deacetylation, as well as the downregulation of HOXC8 expression, are prevented by cyclosporin A and by trichostatin A, suggesting that the prolyl-isomerase activity of Cyp33 is necessary for these effects, and that histone deacetylation mediates these effects. We have previously shown that overexpression of Cyp33 enhances the recruitment of HDAC1 to the repression domain of MLL. We now show that this enhanced recruitment is inhibited by cyclosporin A. These results suggest a model in which Cyp33 is recruited to MLL and targets the MLL repression domain to promote the recruitment of histone deacetylases, thus allowing or promoting the silencing of the MLL regulated genes. In this model Cyp33 functions downstream of the gene activation or silencing epigenetic marks.
A Conserved Regulatory Element in the Mammalian β-Globin Promoters.
Authors: Wada-Kiyama Yuko , Kato Megumi , Onishi Yoshiaki , Kiyama Ryoiti
Published: October 2019

Recent progresses in the study of histone modifications in gene networks and signaling revived the importance of chromatin for controlling gene functions. However, the difficulty of chromatin study lies first in the number of necleosomes in a cell: roughly 107 nucleosomes, each containing a different DNA sequence, are continuously tied as ”beads and strings”, interacting each other, between the neighboring ones as well as the distant ones. Chromatin organization and nucleosome stability is crucial in massive changes of chromatin enviroment such as in DNA replication and formation of chromosomes at cell division as well as in the local changes as observed in transcription of genes, changes of active/inactive chromatin regions and their boundary formation. One of the best ways to study this complex state is first to classify these nucleosomes based on some definitive criteria. Realization that all nucleosomes are not behaving identically is not new. However, explanation at the molecular level of such differences needs a comprehensive analysis of the genome structure by molecular biological experiments and by computational extractions which became available only in recent years. We provide here evidence for a conserved regulatory element for transcription of the β-like globin gene as a summary of our recent studies of a total of over 30 genes using experimental and/or computational approaches (Onishi & Kiyama. Nucl. Acids Res.29, 3448–3457, 2001; Onishi & Kiyama J. Biol. Chem.278, 8163–8171, 2003; Kato et al. J. Mol. Biol.332, 111–125, 2003; Wanapirak et al. J. Mol. Evol.56, 649–657, 2003; Onishi & Kiyama. Recent Res. Devel. Nucl. Acids in press). The element is characterized by the appearance of AA- or TT-dinucleotides in the A+T-rich region located 200 to 400 bp upstream of cap sites. G tracts 3 to 5 nucleotides long exist between the A+T-rich region and the conserved transcription factor binding sites (GATA-1 site and the CACCC, CCAAT, and ATA boxes). An average periodicity of AA- or TT- dinucleotides in the region from a total of eighteen β-family globin genes from four species was approximately 13 bp, suggesting DNA in these regions to show right-handed superhelicity. A proposed biological meaning of this element is to adjust the spatial positions for the first interaction of the transcription factors which should recognize specific DNA sequences in the presence of packed chromatin.
P21 Induces Apoptosis in 32d Cells Undergoing G-Csf-Stimulated Differentiation--This Effect Requires Threonine 57 in Its Cdk2-Inhibitory Domain.
Authors: Ghanem Louis , Steinman Richard A.
Published: October 2019

p21waf1 has been reported to promote hematopoietic stem cell quiescence and to be overexpressed in a subset of leukemias. p21 has been shown to function both as an agonist and antagonist of apoptosis, cell cycling and differentiation in vitro. Functions of p21 and of phosphorylated forms of p21 in granulopoiesis have not been examined hitherto. We hypothesized that p21 would antagonize granulocytic differentiation based on oberved downmodulation of p21 during normal granulopoiesis and the reported antagonism of kerotinocyte differentiation by exogenous p21. The murine myeloblast cell line 32Dcl3, which proliferates in IL-3 and differentiates in G-CSF, was used to model the effects of exogenous wild type and mutant p21 on granulocytic differentiation. Our results demonstrate that the level of p21 protein decreased upon initiation of differentiation. The decrease in p21 protein was secondary to a decrease in p21 message stability as well as proteosomal degradation. Conceivably, the observed decrease in p21 protein was necessary for proper differentiation. In order to determine this, we established stable retroviral transfectants of 32Dcl3 cells to express exogenous wild-type p21, or p21 mutated at threonine-57, threonine-145, serine-146 or both threonine 145 and serine 146 (other studies have shown that mutation of T57 within the cdk2-binding domain of p21 makes p21 resistant to GSK3beta-mediated degradation; the T145 and S146 residues are substrates of Akt which is activated downstream of IL-3 in 32Dcl3 cells). Overexpression of wild-type, T145 or S146 mutants caused growth arrest and triggered apoptosis within 48 hours of stimulation with G-CSF when compared to controls. In contrast, the T57A p21 mutant neither caused growth arrest, nor did it trigger apoptosis in G-CSF. These data support a physiologic role for the observed downmodulation of p21 in differentiaing 32D cells and suggest that p21 is proapoptotic in differentiating granulocytes. The threonine 57 site in the cdk2-binding domain of p21 is implicated in this function.
Rituximab and Stem Cell Transplantation Produces Durable Remissions in Mantle Cell Lymphoma.
Authors: Capote Francisco J. , Pascual M. J. , Gonzalez-Barca E. , Bergua J. M. , Jimenez A. , Romero A. , Garcia-Boyero R. , Giraldo P. , Fernandez-Calvo J. , Ribera J. M. , Ramirez M. J. , Amigo M. L. , Munoz M. I. , Caballero D. , Guinot M. , Oriol A. , Moraleda J. M. , Palomera L. , Almagro M. , Leon A. , Fernandez-deSevilla A. , Gil J. L.
Published: October 2019

Introduction: Mantle cell lymphoma (MCL) is a CD20+ malignancy comprising up 5% of non-Hodgkin’s lymphomas, and has a poor prognosis under standard chemotherapy. The HyperCVAD-M/A regimen (fractionated high-dose cyclophosphamide, vincristine, doxorubicin and prednisolone alternated with methotraxate and cytarabine) has yielded encouraging results when combined with autologous stem cell transplantation (ASCT) in MCL, with 5-year failure-free survival of 54% and overall survival 72%. In an effort to improve these results further, we have combined rituximab in vivo purging and post-transplant consolidation with HyperCVAD-M/A plus ASCT.

Methods: Patients aged <65 years with previously untreated or relapsed MCL were treated with four courses of HyperCVAD-M/A followed by four once-weekly doses of rituximab 375mg/m2 as purging prior to stem cell mobilization and harvesting, high-dose chemotherapy (ICT-CY or BEAM), stem cell reinfusion and four further doses of rituximab immunotherapy post-transplant.

Results: Of the 34 patients enrolled so far, 15 (12 male, 3 female; 12 previously untreated) have been transplanted. The median age was 52 years (range 47–63 years). After the final post-ASCT immunotherapy all 15 patients were in clinical complete remission. With a median follow-up of 30 months from diagnosis (range 7–52 months), 14 patients remain alive with 13 in first complete remission. One patient died 15 months post-ASCT without evidence of disease recurrence. Kaplan-Meier estimates of 4-year overall and event-free survival are 93.3% and 86.6% respectively.

Conclusions: This approach seems safe and feasible and produces durable remissions; longer follow-up of a more patients will be required to assess the effect of the procedure on survival.
The Use of Gene-Expression Profiling to Predict Outcome of Follicular Lymphoma Patients Treated with Rituximab and Chemotherapy.
Authors: Harjunpää Antti , Nykter Matti , Monni Outi , Karjalainen-Lindsberg Marja-Liisa , Hemmer Samuli , Yli-Harja Olli , Meri Seppo , Leppä Sirpa
Published: October 2019

The anti-CD20 monoclonal antibody rituximab has been extensively evaluated for treatment of follicular lymphoma (FL) and is now an integral component of many treatment strategies. However, it is not known when and how to use this combination modality best in the overall treatment course of FL patients. Here we have analyzed the pattern of gene expression in lymph nodes from 28 FL patients who received rituximab in combination with chemotherapy. We demonstrate a novel feature selection method, in which genes whose expression correlates with treatment outcome are devided into groups based on similar expression patterns. By selecting representatives from these groups we built a three-gene predictor (for example RRAD, MGC5254, MARCO) which turned out to be successful in classifying all 28 samples correctly with respect to the responders versus nonresponders distinction. A similar four-gene predictor (for example USP9Y, CUL-4B, NSFS/REST, MADH) correctly classified the responding patients into two subgroups with very different event-free survival rates at the median follow up of 21 months (median not reached vs. 15 months, p<0.0001). The latter predictor also efficiently delineated patients within specific FLIPI risk groups. Our data demonstrate the ability of these microarray-based predictors to classify FL patients treated with rituximab and chemotherapy to subgroups with significantly distinct outcomes.
Characterization of Cellular Immune Responses to a Recombinant Idiotype Vaccine by ELISPOT and Identification of MHC Class I-Restricted T Cell Epitopes by Peptide Mapping.
Authors: Bertinetti Cristina M. , Veelken Hendrik
Published: October 2019

Induction of tumor-specific immune responses by idiotype vaccination is a promising strategy for biological therapy of indolent B cell lymphomas. In a previous report, we have described immune responses in a subset of patients participating in a phase I clinical trial primarily designed to demonstrate safety and efficacy of a recombinant idiotype vaccine (Veelken et al., ASH abstract #3342, 2003). In this trial, B-NHL patients who had relapsed after standard chemotherapy received repetitive intradermal vaccinations with recombinant idiotype Fab fragment derived from their tumor mixed with lipid-based adjuvant and concurrent subcutaneous GM-CSF at the same site. We now present the final analysis of cellular immune responses in this cohort. Peripheral blood lymphocytes (PBL) were obtained prior to and on various time points during and after vaccinations. Cryopreserved PBL were stimulated twice by autologous dendritic cells (DC) exposed to the autologous Fab protein for cross-presentation as MHC class I-bound peptides. INFγ-secreting cells were subsequently quantified by ELISPOT with Fab-presenting DC. Alternatively, freshly thawed PBL were directly assayed with recombinant Fab by ELISPOT without prestimulation. An increase in the frequency of Fab-responding PBL was detected in 7 of 15 evaluable patients with the prestimulation assay and in 4 of 10 patients by direct quantitation, resulting in a combined cellular response rate of 53% (9 of 17). A cellular immune response showed a trend for correlation with extended progression-free survival (p=0.08). T cell responses were predominantly idiotype-specific since lesser or no increases in IFNγ-secreting cells were detected against light chain- and VH family-matched control Fabs. Interestingly, a much higher base-line reactivity was observed against the control Fabs in comparison to the patient’s lymphoma Fab in four patients, pointing to the possibility of tumor-specific anergy in lymphoma patients that can at least be partially corrected by active immunization. In an effort to identify the MHC class I-presented idiotype-derived peptides, potential binding motifs were defined by reverse immunology with the SYFPEITHI algorithm (www.syfpeithi.de). Ten candidate peptides from the variable and constant region of an immune responder’s idiotype heavy chain were synthesized and evaluated with post-vaccination PBL by ELISPOT without prestimulation. A peptide derived from the CDR2 region showed a significantly higher response compared to an unrelated peptide control (p=0.0013). Additional peptides derived from the FWR1, CDR1, and CDR2 also showed a significant stimulation, but only in comparison to a no peptide control. ELISPOT offers a valuable tool to monitor cellular immune reponses and demonstrates successful induction of tumor immunity in pretreated, tumor bearing and immunosuppressed B cell lymphoma patients.

Supported by Deutsche Krebshilfe
High Dose Therapy with Autologous Blood Stem Cell Transplantation (Asct) for Relapsed, Follicular Lymphoma (FL) vs. De Novo, Diffuse Large B-Cell (dnDLBL) vs. Transformed, Follicular To Diffuse Large B-Cell Lymphoma (Tdlbl): No Difference in Freedom from Progression.
Authors: Pohlman Brad , Jin Tony , Summers Kristie , Kuczkowski Elizabeth , Brown Stacey , Kalaycio Matt , Sobecks Ronald , Andresen Steven , Bernhard Laura , Cherni Kelly , Baker Janet , Bolwell Brian J.
Published: October 2019

The decision to offer or not offer high dose therapy with ASCT to an individual lymphoma patient is sometimes (but perhaps mistakenly) based on the specific histology alone and not necessarily on patient characteristics, clinical manifestations, or disease behavior. Therefore, we reviewed the long-term outcome of FL, dnDLBL, and tDLBL patients, who received high dose chemotherapy (CT) with ASCT at the Cleveland Clinic, to determine if the specific histology is important. Between June 1991 and July 2004, 235 patients with FL, dnDLBL, or tDLBL in second or third remission received high dose CBV (n=7), BuCy (n=1), or BuCyVP (n=227) with ASCT. The median follow-up among survivors is 3.4 (.1–11.4) years.

Patient, disease, and ASCT characteristics and outcome according to histology

Variable FL (n=88) dnDLBL (n=123) tDLBL (n=24) p-value Age: median (range) 51(33–69) 50(22–71) 56(40–70) 0.021 Male sex: N (%) 47(53) 73(59) 12(50) 0.56 Years from diagnosis to ASCT: median (range) 2.6(0.4–17.5) 1.5(0.3–15.6) 3.4(1–14.0) <0.001 Prior #CT regimens: median (range) 2(1–5) 2(1–4) 2(2–5) <0.001 Disease status at ASCT: CR2-PR2/CR3-PR3, N (%) 61(69)/27(31) 106(86)/17(14) 16(67)/8(33) 0.005 Bone marrow involved at ASCT: N (%) 16(21) 12(10) 1(4) 0.035 Prior radiation therapy: N (%) 25(28) 45(37) 7(29) 0.42 LDH > normal at ASCT: N (%) 60(69) 65(54) 12(50) 0.06 Tumor bulk >10 cm at ASCT: N (%) 14(16) 27(23) 4(17) 0.46 Disease progression: N (%) 35(40) 55(45) 10(42) – Death from lymphoma: N (%) 21(24) 42(34) 7(29) – Death from any cause: N (%) 31(35) 58(47) 13(54) –

Kaplan-Meier freedom from progression curves accarding to histology are shown:

Figure Figure

Kaplan-Meier overall survival curves according to histology are shown:

Figure Figure

There is no significant difference in freedom from progression between FL, dnDLBL, and tDLBL patients transplanted in second or third remission. By Cox proportional univariate analysis, only male sex predicted a higher risk of progression while male sex, older age, and dnDLBL or tDLBL (compared to FL) predicted a higher risk of death. By Cox proportional multivariate analysis, no factor predicted a higher risk of progression while older age, male sex, and dnDLBL predicted a higher risk of death. In conclusion, high dose chemotherapy with ASCT leads to long-term remissions in 40–50% of FL, dnDLBL, and tDLBL patients in second or third remission. These results suggest that the distinction between these three histologies is less important than other factors in determining patient eligibility for ASCT.
Ligand Activation of FLT3 Induces Cell Cycle Arrest in Acute Lymphoblastic Leukemia with 11q23 Translocation.
Authors: Furuichi Yoshiyuki , Sugita Kanji , Inukai Takeshi , Goi Kumiko , Takahashi Kazuya , Nakazawa Shinpei
Published: October 2019

A fms-like kinase (FLT3) is widely known to be involved in proliferation of normal hematopoietic stem cells and precursors. Acute lymphoblastic leukemia (ALL) with 11q23 translocation, often found in infantile leukemia with very poor prognosis, is thought to be derived from a population of cells at the developmental stage very close to hematopoietic stem cells, and was recently shown that they express FLT3 at high levels compared with other types of leukemia. In the present study, we examined the effects of FLT3 ligand (FL) on leukemia cells with or without 11q23 translocation to evaluate a biological implication of the FLT3/FL interaction in ALL. Three of 8 leukemic cell lines without 11q23 translocation showed a proliferative response to FL in the 3H-thymidine uptake assays. However, five of 7 B-precursor leukemic cell lines with 11q23 translocation, unexpectedly, showed an inhibitory response (23-69% inhibition) to FL in a dose-dependent manner (1–20ng/ml), although the special cell line with D835 mutation in FLT3 (KOCL-33) was not affected by the addition of FL. This inhibitory effect was almost abrogated in the presence of a FLT-3 kinase inhibitor PKC412. Inhibition of 3H-thymidine uptakes were not due to induction of apoptosis but due to induction of the Go/G1 arrest. This cell cycle arrest was mediated, at least in part, by a marked up-regulation of p27 due to suppression of its degradation, and promoted resistance of cell lines to radiation-induced apoptosis. Of interest, the addition of FL induced a complete disappearance of constitutive phosphorylation of STAT5 but upregulated phosphorylation of MAPK and Akt. These results suggest that the FLT3/FL interaction in ALL with 11q23 translocation transmits the inhibitory signal specifically to the JAK/STAT pathway via the kinase activity of FLT3, in the process of which the JAK/STAT-specific inhibitory molecules such as SOCS-2 and CIS-1 may be implicated.
A Role for Adipocyte-Enriched Transcription Factor, PPAR-γ, in Up-Regulation of PAI-1 Gene Expression during Adipogenesis.
Authors: Ihara Hayato , Loskutoff David J. , Urano Tetsumei
Published: October 2019

Obese patients are at risk for development of cardiovascular disease, which can in part be explained by disturbances in the haemostatic and fibrinolytic systems. Recently, it has been demonstrated that the adipocyte itself is able to produce a primary fibrinolytic inhibitor, plasminogen activator inhibitor-1 (PAI-1), possibly explaining the high levels found in obesity. We have previously shown that pioglitazone, a selective ligand for peroxisome proliferator-activated receptor (PPAR)-γ, and insulin not only enhances adipocyte differentiation, but also up-regulates PAI-1 gene transcription in 3T3-L1 adipocytes (Ihara et al. FASEB J., 2001). These results suggest that PAI-1 gene expression during adipogenesis can be regulated by a direct action of PPAR-γ. In the present study, we investigate a role of an adipocyte-enriched transcription factor, PPAR-γ, in up-regulation of PAI-1 gene expression in adipose tissues. To identify cis-acting genetic elements required for induction of PAI-1 gene transcription, we constructed the plasmids containing a 5′ deletion series of the PAI-1 promoter. When sequences between -762 and -723 were deleted, the level of induction significantly fell. By sequence homology search in this region, we found an atypical PPAR-responsive element (aPPRE) at positions −752 to −733. Gel mobility shift and supershift assays using oligonucleotides of the aPPRE overlapping C/EBP binding site indicated that the aPPRE is important for PPAR-γ binding. Chromatin immunoprecipitation assay demonstrated that PPAR-γ is physically associated with PAI-1 gene promoter region including the aPPRE in vivo. Furthermore, mutation of the aPPRE abrogated the increase in transcriptional activities of PAI-1 promoter. Functional assay revealed that besides PPAR-γ/RXR-α expression vectors, co-transfection of CREB-binding protein expression vector increased the luciferase activity to 7.5 fold and 11 fold of control value in COS-1 cells and 3T3-L1 preadipocytes, respectively. These data indicate that the transcriptional induction of PAI-1 gene during adipogenesis is mediated through the binding of PPAR-γ/RXR-α to the functional aPPRE in the PAI-1 gene promoters.
Molecular Mechanism of Prothrombin G20210A Variant: Critical New Role of Exon Splicing Enhancer in Poly (A) Tailing.
Authors: Kuwahara Mitsuhiro , Kurachi Sumiko , Kurachi Kotoku
Published: October 2019

Although prothrombin G20210A variant was first found in 1996 as a genetic risk factor of thrombosis, the underlying molecular mechanism for elevation of circulatory prothrombin levels due to the variant has remained elusive. The position 20210 is at the poly (A) tailing site, and several research groups reported different, contradictive possibilities including enhancement in poly (A) tailing, change in RNA stability or no difference in prothrombin mRNA level between G (wild type) and A (variant) at position 20210. Such confusions appear to be due to no reliable assay model availability. To overcome these problems, we developed a minigene assay system composed of only autologous components of human prothrombin gene, including its promoter, coding regions, introns and 3′UTR with poly (A) signal.

A minigene containing the first intron and the last intron did not express at any appreciable levels, failing proper splicing of the last intron. Inclusion of upstream other introns was found essential for its appropriate splicing. Although it is generally accepted that the last intron enhances the efficiency of poly (A) tailing, we found that it is not the case for the prothrombin gene. Instead, an exon-splicing enhancer (ESE) of another upstream intron was found to be responsible for efficient poly (A) tailing. ESE, therefore, plays a critical role not only in alternative splicing but also in constitutive RNA processing, including poly (A) tailing. Etiology of prothrombin G20210A variant studied with culture cells and transgenic mice, supported an enhancement of poly (A) tailing, and not a change in mRNA stability.
Genetic Polymorphisms in the Promoter Region of the beta-2 Adrenergic Receptor Are Associated with the Early Response of Acute Lymphoblastic Leukemia to Chemotherapy.
Authors: Cheok Meyling H. , Ding Cong , Yang Wenjian , Das Somas , Campana Dario , Cheng Cheng , Cook Edwin H. , Pui Ching-Hon , Relling Mary V. , Evans William E.
Published: October 2019

Acute lymphoblastic leukemia (ALL) in children is a paradigm of disseminated cancer that is curable with chemotherapy, yet current treatment fails to cure about 20% of patients, for reasons that remain unknown. In a genome-wide assessment of in vivo treatment-induced changes in gene expression in ALL cells using the Affymetrix U95A and U133A oligonucleotide microarray, we found that patients who eventually relapsed did not up-regulate expression of the pro-apoptotic beta-2 adrenergic receptor gene (ADRB2) in their ALL cells after initial treatment with methotrexate and mercaptopurine. After treatment we observed a 5-fold lower level of ADRB2 gene expression in leukemia cells of patients who ultimately relapsed. We found a common genetic polymorphism in the ADRB2 promoter that was significantly linked to high-dose methotrexate induced up-regulation in ADRB2 gene expression in ALL cells. Moreover, the ADRB2 promoter haplotype was significantly linked to poor early treatment response in ALL cells from 242 children (i.e., probability of event-free survival at two years, p=0.0275 stratified by risk groups). These findings have revealed a germline polymorphism that is linked to the early antileukemic effects of ALL chemotherapy and provide new insights into genetic determinants of ALL treatment efficacy.
Bone Mineral Density, Fracture Risk and Avascular Bone Necrosis in Childhood Acute Lymphoblastic Leukemia; an Up-Date of the Prospective DUTCH ALL9 Study.
Authors: Van den Heuvel-Eibrink Marry M. , Van der Sluis Inge M. , Leeuw Bert A. , Kardos Gaby , te Winkel Lizet M. , Revesz Tom , Korthof Elisabeth T. , Bokkerink Jos , Veerman Anjo J.P.
Published: October 2019

The high cumulative dose of dexamethasone, applied in the DCOG ALL9 protocol, prompted us to investigate the risk of osteoporosis, fractures and avascular necroses of bone (AVN) in children treated with acute lymphoblastic leukemia (ALL). Fracture risk and incidence of symptomatic AVN was assessed in 778 patients(482 boys, 297 girls), included in the ALL9 protocol since 1997. Total cumulative doses (TCD) of dexamethasone were 1370 mg/m2 and 1244 mg/m2 and of MTX 8.1g/m2 13.6g/m2 for NHR and HR patients respectively. No CNS-irradiation was applied. In children aged >3 years, lumbar spine bone mineral density (BMD) was measured by dual-energy X-ray absorptiometry (DEXAscan), at diagnosis(T0), after 32 weeks(T1), at discontinuation of treatment at 109 weeks(T2), and one year after discontinuation of treatment(T3). Results were expressed as standard deviation scores (SDS). Symptomatic AVN was defined as on MRI confirmed AVN lesions in combination with non-vincristine related persistent pain in arms or legs. Fractures were reported in 82/778 (10.5%) patients. Most occurred after mild trauma. No difference was found in fracture incidence between boys and girls. BMD was measured in 387/427 (90.6%) eligible patients. Median BMD-SDS was significantly lower than zero at all times of evaluation, the lowest BMD values were found at T2 (−1.47 SDS). Fracture risk was 3.9 times higher as compared to healthy school children. Fracture incidence was correlated with BMD at T2 and T3(p=0.04 and p=0,04 respectively), but not at T0 and T1. A significant more rapid decline in BMD from T0 to T2 and to T3 was seen in patients with fractures as compared to patients without fractures. After discontinuation of therapy, BMD recovered faster in cases without fractures. Symptomatic AVN occurred in 33/778 (4.2%) of our patients (med age 14, range 6,5–18 years) showing irreversibility in 22 % of the cases. Differences found in the incidence between the centers may suggest underestimation of the risk of fractures and AVN in this prospective study. Children with ALL show a significantly increased fracture risk. Patients with a more severe reduction in BMD during treatment are more susceptible to fractures. The AVN incidence in this protocol did not exceed previous reports of prednisolone-based protocols.
The Heme Oxygenase-1-Targeting Compound PEG-ZnPP Inhibits Growth of Imatinib-Resistant BCR/ABL-Transformed Cells.
Authors: Mayerhofer Matthias , Aichberger Karl J. , Florian Stefan , Krauth Maria-Theresa , Derdak Sophia , Esterbauer Harald , Wagner Oswald , Pickl Winfried F. , Selzer Edgar , Deininger Michael W. , Druker Brian J. , Greish Khaled , Maeda Hiroshi , Sillaber Christian , Valent Peter
Published: October 2019

Chronic myeloid leukemia (CML) is a myeloproliferative disease characterized by the BCR/ABL oncogene and an increased survival of leukemic cells. The BCR-ABL tyrosine kinase inhibitor imatinib has successfully been introduced as a treatment of CML. However, resistance after an intitial response is common in patients with advanced disease, and it is not yet clear if responses in early disease phases will be durable. Therefore, current studies focus on novel potential drug-targets in CML cells. We have recently identified heme oxygenase-1 (HO-1) as a novel BCR/ABL-dependent survival-molecule in primary CML cells. In this study, we analyzed signal transduction pathways underlying BCR/ABL-induced expression of HO-1 and evaluated the role of HO-1 as a potential new target of drug therapy. We found that the PI3-kinase inhibitor LY294002 and MEK inhibitor PD98059 downregulate expression of HO-1 in CML cells. In addition, constitutively active Ras- and Akt -mutants were found to promote expression of HO-1 in Ba/F3 cells, further supporting the involvement of the PI3-kinase/Akt as well as the MAPK pathway in regulating HO-1 expression. To establish a role for HO-1 in survival of CML cells, expression of HO-1 was silenced by siRNAs which resulted in apoptosis of K562 cells. Next, HO-1 was targeted in CML cells by pegylated zinc protoporphyrin (PEG-ZnPP), a competitive inhibitor of HO-1. Exposure to PEG-ZnPP resulted in growth inhibition and induction of apoptosis in primary CML cells as well as in the CML-derived cell lines K562 and KU812 with IC50 values ranging between 1–10 μM. The growth-inhibitory effects of PEG-ZnPP were not only observed in CML cells responsive to imatinib, but also in imatinib-resistant K562 cells and Ba/F3 cells expressing various imatinib-resistant mutants of BCR/ABL (T315I, E255K, M351T, Y253F, Q252H, H396P). Moreover, imatinib and PEG-ZnPP were found to exert synergistic growth inhibitory effects on imatinib-resistant leukemic cells. Together, these data suggest that HO-1 represents a novel drug target in cells expressing BCR/ABL, including those with resistance to imatinib.
High Levels of CD34 Expression in Bone Marrow Trephines Predicts for Loss of Cytogenetic Response in Patients with in Chronic Phase Who Have Achieved Complete Cytogenetic Remission on Imatinib Therapy.
Authors: Marin David , Horncastle D. , Howard Jill , Andreasson Catherina , Elliot V. , Goldman Jonh M. , Apperley Jane F. , Naresh Kikkeri , Lampert Irvin
Published: October 2019

We studied the expression of CD34 in bone marrow trephines in 71 patients with CML in CP who had achieved complete cytogenetic remission (CCyR) on imatinib in order to predict the durability of the cytogenetic response. 26 patients received the imatinib shortly after diagnosis while 45 were treated after interferon failure. Imatinib was given at a dose of 400 mg daily. Imatinib dosage was adjusted depending on tolerance and response. Bone marrow examinations were performed every 3-6 months. The CD34 stain was performed in paraffin sections with the monoclonal antibody QBEND10. A minimum of 500 nucleated cells were counted per trephine section. Results were expressed as percentage of CD34+ vs all nucleated marrow cells. The expression of CD34 by the endothelial cells was used as an internal positive control. The median follow up was 1120 days (percentile 25 and 75: 960 and 1254 days). The median time to the achievement of CCyR was 254 days (percentile 25 and 75: 167 and 380 days). Eight patients lost CCyR at a median time of 981 days (range 546 to 1428 days). For the study we used the first evaluable bone marrow taken after the start of the imatinib therapy (median time 242 days; percentile 25 and 75: 110 and 342 days. The median value of CD34 was 0.9% (percentile 25 and 75 percentiles: 0.8 and 1%). Univariate and multivariate analysis for loss of CCyR was performed. The variables included in the analysis were: age, sex, Sokal and Euro risk groups, percentage of blasts in the bone marrow aspirate, new patient vs IFN intolerant, time from diagnosis to the start of the imatinib therapy and the presence of additional cytogenetic abnormalities. The only variable found to be significant in the univariate and in the multivariate analysis was the level of CD34, RR=2.45 (95CI 1.29–4.64, p=0.006). When the variable was categorised as CD34 level <=2% and CD34 level >2% the cumulative incidence of loss of CCyR at 4 years was 10.58% (95CI 3.8–10.6%) in the patients with a CD34 <=2% (n=64) vs 75% (95CI 31.9–95.1%) in the patients with a CD34 >2% (n=7), p<0.0001. BCR-ABL positive hematopoietic progenitors resistant to imatinib are detectable in all the patients who have achieved cytogenetic remission (Bhatia, 2003. Graham 2002) and may be important in disease relapse. We have shown that patients in CCyR whose percentage of CD34 positivity in trephine biopsies. taken shortly after commencing imatinib, is in excess of 2% are much more likely to lose cytogenetic remission than those with lower levels. We speculate that an increase in CD34 expression could reflect an increase in leukemic precursors in some patients.
Clinical Useful Prognostic Index for Adult Patients with Acute Myeloid Leukemia in First Relapse.
Authors: Breems Dimitri A. , Van Putten Wim L.J. , Lowenberg Bob
Published: October 2019

The treatment of acute myeloid leukemia (AML) in first relapse is associated with unsatisfactory rates of complete responses that are usually short lived. Previously proposed prognostic classification methods serving therapeutic decisions and evaluation of investigational treatment strategies at relapse of AML have been based on the duration of the relapse free interval and have largely neglected the influence of other known prognostic factors. Here we present an improved clinically useful prognostic index. This index has been developed from a multivariate analysis of 667 AML patients in first relapse among 1540 newly diagnosed non-M3 AML patients of age 15 to 60 years entered into three successive HOVON/SAKK Collaborative Group trials. The score, which has a range of 0 to 14 points, uses four relevant parameters. The parameters are: length of relapse free interval after first complete remission (more than 18 months: 0 points; 7 to 18 months: 3 points; 6 months or less: 5 points), cytogenetics at diagnosis (t(16;16) or inv(16): 0 points; t(8;21): 3 points; other cytogenetics: 5 points), age at relapse (35 years or younger: 0 points; 36 to 45 years: 1 point; older than 45 years: 2 points) and whether or not a previous stem cell transplantation (SCT) has been undertaken in first complete remission (no SCT: 0 points; previous SCT: 2 points). These points were assigned following estimations of the relative values of each of these factors contributing to outcome. Ultimately, three risk groups were defined: a favorable prognostic group A (0 to 6 points; overall survival of 70% at one year and 46% at five years), an intermediate risk group B (7 to 9 points; overall survival of 49% at one year and 18% at five years), and an unfavorable risk group C (10 to 14 points; overall survival of 16% at one year and 4% at five years). Thus, four commonly applied clinical parameters may identify among patients with AML in first relapse those for salvage or investigational therapy.