bluuebunny/crossref_metadata_2025_split huggingface dataset:
Once Per Cycle Pegfilgrastim Versus Daily Filgrastim in Pediatric Patients with Ewing Sarcoma.
Authors: Wendelin Gerald , Lackner Herwig , Schwinger Wolfgang , Sovinz Petra , Urban Christian 
Published: October 2019

Abstract
The administration of the recombinant human granulocyte colony-stimulating factor (rhG-CSF) Filgrastim for reducing the duration of severe neutropenia after cytotoxic chemotherapy has become an important part of oncologic supportive care. Due to the short serum half-life Filgrastim has to be administrated daily by subcutaneous injections. Frequent injections however mark a problem in pediatric patients. Studies in adult patients have shown a comparable effect of the new long lasting rhG-CSF Pegfilgrastim which has to be administrated only once per cycle. In the current study the effects of Pegfilgrastim in pediatric patients were analysed. Five patients (10–16 years) with Ewing sarcoma were treated in a cross over study design alternately with Pegfilgrastim and Filgrastim following the EURO E.W.I.N.G. 99 protocol. Starting on day 4 after chemotherapy patients received Filgrastim 10μg/kg daily by subcutaneous injection until an absolute neutrophil count (ANC) >1000/μl after the expected nadir. Pegfilgrastim 100μg/kg was administrated on day 4 once per cycle subcutaneously. In 3 patients the stimulation with rhG-CSF was performed after each of the 6 preoperative VIDE-(Vincristin, Ifosfamide, Doxorubicin, Etoposide) cycles, in 2 patients after 8 postoperative VAI-(Vincristin, Actinomycin D, Ifosfamide), and in 2 patients after 7 postoperative VAC-(Vincristin, Actinomycin D, Cyclophosphamide) cycles of the EURO-E.W.I.N.G. 99 protocol. The duration of grade 4 neutropenia after single administration of Pegfilgrastim was 2,8 ±3,1 (0–10) days, after daily administration of Filgrastim 3,1 ± 2,7 (0–8) days. The number of days with a body temperature over 38 degrees and grade 4 neutropenia at the same time was 0,9 ±1,5 (0–6) after Pegfilgrastim and 0,9± 1,4 (0–4) after Filgrastim. Filgrastim had to be injected 6,7 ± 1,8 (3–10) times per cycle. Bone pain associated with Pegfilgrastim was noted in only one patient. Costs for Pegfilgrastim were 16% lower than for Filgrastim. We conclude that in pediatric patients with Ewing sarcoma the duration of severe neutropenia and number of days with febrile neutropenia after once per cycle Pegfilgrastim and daily Filgrastim are comparable. By using Pegfilgrastim the number of subcutaneous injections can be reduced to one single injection and costs can be lowered.
Interferons and IRF8/ICSBP Stimulate Immune Protection Against BCR/ABL-Induced Leukemia.
Authors: Nardi Valentina , Mohammad Azam , Dikovskaia Eugenia , Daley George Q. 
Published: October 2019

Abstract
Chronic Myeloid Leukemia (CML) results from transformation of hematopoietic stem cells by the BCR/ABL oncoprotein. CML can be effectively treated with the targeted ABL kinase inhibitor Imatinib (STI571; Gleevec), but patients rarely achieve molecular remission by sensitive PCR methods, and drug resistance limits the durability of drug response. A significant minority of patients treated with Interferon (IFN) alpha achieve complete cytogenetic remission, and response is correlated with immune reactivity against leukemic clones. This suggests that IFN-induced immunotherapy might be an important adjunct to Gleevec, thus prompting us to investigate immunoprotective mechanisms. We have shown previously that ectopic expression of Interferon Regulatory factor 8 (IRF8, also called ICSBP) can stimulate a potent CD8+ cytotoxic anti-leukemia response in a murine model of BCR/ABL-induced leukemia (Deng and Daley, Blood 97;3491, 2001). In transcriptional profiling experiments with cells that overexpress IRF8/ICSBP, we detected increased expression of IFN alpha and beta. To test whether IFN alpha or beta can mediate the same protective immune-response as IRF8/ICSBP, we co-injected BCR/ABL-transformed BaF3 cells together with non-transformed BaF3 cells engineered to overexpress IFN alpha or beta. Expression of IFN alpha or beta in trans protected mice from developing leukemia. Control constructs in which IFN alpha or beta were cloned in the anti-sense orientation failed to confer protection, confirming the importance of IFN alpha or beta expression. Importantly, IRF8/ICSBP expression is protective against cells transformed by Gleevec-resistant mutant BCR/ABL proteins (T315I and L248R). Our data suggest that IFNs are functionally equivalent to IRF8/ICSBP in conferring a vaccine-like anti-leukemic immune effect in our model, and may act through an autoregulatory loop involving IRF8/ICSBP. Defining the precise epitopes targeted by the IFN/ICSBP-induced immune response may enable immunotherapy in human CML patients.
A New Functional Coagulation Assay for Measurement of Total Factor V Activity in Human Plasma and Its Application to Samples from Patients with Disseminated Intravascular Coagulation.
Authors: Samis John A. , Stewart Kenneth A. , Toh Cheng Hock , Downey Colin , Nesheim Michael E. 
Published: October 2019

Abstract
Factor V (F.V) circulates as an inactive single chain procofactor. Only upon cleavage and activation with thrombin to F.Va does it function as a cofactor in the F.Xa-mediated conversion of prothrombin to thrombin. The conventional F.V coagulation assay measures the one-stage clotting activity, which most likely reflects the trace of F.V that is activated in the sample or becomes activated during the assay. Since F.V is activated by thrombin to F.Va, a second (two-stage) assay is crucial to fully assess the total F.V potential activity in the plasma sample. For this assay, normal human pooled plasma (diluted 100- to 500-fold) was treated with thrombin (1–5 NIH Units/ml, final concentration) for various times at 37°C to fully activate the F.V. The added thrombin was inhibited with a 2-fold molar excess of Phe-Pro-Arg chloromethylketone (PPACK) and the sample re-assayed. This second assay measures the total F.V activity of a sample. Maximal 10-fold activation of plasma F.V activity occurred with 2Units/ml thrombin after 1 to 2min at 37°C and this activity was stable for 30min prior to PPACK addition. Control experiments indicated that the levels of PPACK used did not affect the one-stage F.V activity of untreated plasma samples but were required to inhibit the added thrombin from clotting fibrinogen in the assay. Assay of timed plasma samples (n=48) from seven patients admitted to the Intensive Care Unit and subsequently diagnosed with disseminated intravascular coagulation (DIC), indicated that the total F.V activity, the specific activity (total F.V activity/antigen), and the activation quotient were decreased, on average, by 45%, 49%, and 31%, respectively, from that observed with normal human pooled plasma. The results indicate that the total amount of activatable F.V as well as its degree of activation decreases from normal as part of the pathogenesis of DIC in humans. Further, the decreased total F.V specific activity observed in DIC patient plasma samples indicates that the F.V that is present is less active per molecule than normal, possibly because of proteolytic inactivation. This total F.V clotting assay should provide new information about the biochemistry of F.V and its contribution to the pathogenesis of coagulation disorders.
Minimal Residual Disease (MRD) Analysis in Acute Myeloid Leukemia (AML) with Favorable Cytogenetics [t(8;21) and inv(16)] by Real Time PCR(RT-PCR) and Flow Cytometry (FC).
Authors: Perea Granada , Lasa Adriana , Aventin Anna , Domingo Alicia , Villamor Neus , de LLano Maria Paz Queipo , Llorente Andreu , Junca Jordi , Palacios Carlos , Fernandez Cristalina , Guallart Miquel , Font Llorens , Tormo Mar , Florensa Lourdes , Bargay Joan , Marti Josep M. , Vivancos Pilar , Torres Pio , Berlanga Juan , Brunet Salut , Sierra Jorge , Nomdedeu Josep F. 
Published: October 2019

Abstract
Objectives: To analyze MRD in 65 patients (pts) with good prognosis AML: 30 t(8;21) and 35 inv(16), using both FC and RT-PCR, and to investigate the prognostic value of MRD in the pts outcome.
Methods: MRD was monitored in CR pts (n=55) by FC in 101 follow-up samples obtained after various cycles of treatment, as follows: 40 post-induction (ind), 30 post-intensification (int) and 31 at the end of treatment (ttm), and by RT-PCR in 76 samples: 31, 23 and 22, respectively. In 35 pts the two techniques were applied at the same time of the ttm. MRD by FC was assessed using fixed combinations of three monoclonal antibodies. AML1/ETO and CBFb/MYH11 were analyzed following the BIOMED protocol.
Results: Twenty-seven percent (n=15) of CR pts relapsed: 6 with t(8;21) and 9 with inv(16). The mean MRD by FC was 1.1% after ind, 0.2% after int and 0.1% at the end of ttm. At the end of ttm, the MRD detected by FC in relapsed and not relapsed pts were significativaly different: 0.3% vs 0.08% (p=0.002). By RT-PCR, the mean of fusion transcript copies/ablx104 differed between relapsed and nonrelapsed pts: 2385 vs 122 (p=0.001) after ind, 56 vs 7.6 after int (p=0.0001) and 75 vs 3.3 (p=0.0001) at the end of ttm. Relapses were more commonly observed in those pts with FC MRD level >0.1% at the end of ttm than in pts with ≤0.1%: 50% vs 12% (p=ns); likewise, using RT-PCR, a cutoff level of >10 copies at the end of ttm correlated with high risk of relapse: 80% of pts with RT-PCR >10 relapsed compared to 12% of pts with levels <10 (p=0.009). The overall survival (OS) probability was 86% for pts with CF MRD ≤0.1 at the end of ttm and 0% for pts with MRD >0.1 (p=0.1) and the leukemia free survival (LFS) was 78% and 44%, respectively (p=0.05). For pts with RT-PCR ≤10 at the end of ttm, the OS was 100% and for pts with RT-PCR >10 it was 30% (p=0.007) and the LFS was 87% and 20%, respectively (p=0.001). MRD was identified after ind in 55% of relapsed pts and at the end of ttm in 83% of relapsed pts. Only 1 pt (1/13) with FC MRD <0.1 and RT-PCR <10 at the end of ttm relapsed. For patients in complete remission, the mean copy level of chimeric transcript was higher for pts with t(8;21) than for those with inv(16): 30.2 vs 17.4 (p=0.0001).
Comments: In tandem analysis of MRD by FC and RT-PCR could improve MRD detection in AML pts.
Identification of Natural Killer Cell Receptor Phenotypes Associated with Leukemia.
Authors: Verheyden Sonja J. , Bernier Michel , Demanet Christian J. 
Published: October 2019

Abstract
Introduction: Natural Killer (NK) cells play a key role in defense against tumor cells that have the capacity to downregulate Human Leukocyte Antigen (HLA) class I expression. It has been reported that leukemic cells can have down-regulated expression of HLA class I molecules. Apparently, the NK cells of these patients are not able to destroy these leukemic cells and may allow malignant cells to escape from innate immune control. This failure may be due to the fact that NK cells are part of the malignant clone and therefore might have a decreased function. An alternative hypothesis could be that these patients may display a NK cell Receptor (NKR) genotype incapable of destroying leukemic cells with aberrant expression of HLA class I molecules. The polymorphic nature of the NKR genes generates diverse repertoires in the human population, which display specificity in the innate immune response.
Materials and Methods: In the present study, 11 Killer cell Immunoglobulin-like Receptor (KIRs) and 2 CD94/NKG2 receptors were genotyped by PCR-SSP in 96 leukemic patients and 148 healthy Caucasians.
Results and Conclusion: We report a significant increased frequency of the more inhibitory AB KIR phenotype in leukemic patients compared to the controls (31.1% in healthy controls vs. 51.0% in leukemic patients, Pc = 0.002), which is related to the high prevalence of the inhibitory KIR2DL2 in this population (Pc = 0.007). Moreover, two specific KIR phenotypes AB1 and AB9, including all inhibitory KIRs, were significantly associated with leukemic patients. Our study suggests that an important percentage of leukemic patients express a KIR phenotype in favor of escape from NK cell immunity.
Phenotype Correction of Hemophilia A Mice with Adeno-Associated Virus (AAV) Vectors Carrying the B Domain Deleted Canine Factor VIII Gene.
Authors: Mimuro Jun , Ishiwata Akira , Kashiwakura Yuji , Takano Katsuhiro , Ohmori Tsukasa , Madoiwa Seiji , Mizukami Hiroaki , Ozawa Keiya , Sakata Yoichi 
Published: October 2019

Abstract
Hemophilia A is a life-threatening bleeding disorder caused by mutations in the factor VIII (FVIII) gene that lead to deficiency of FVIII. Gene therapy is respected to provide an alternative to current FVIII supplemental therapy. Among a variety of vectors, AAV vectors are thought to be ideal for transfer of therapeutic genes since they are derived from non-pathogenic virus and have been demonstrated to provide sustained transgene expression in non-dividing cells with little toxicity, though, delivery of the FVIII gene using AAV vectors are limited by its small packaging capacity. To overcome the packaging capacity limit, we developed AAV1 vectors and AAV8 vectors carrying the B domain deleted canine FVIII gene (BDD cFVIII) utilizing a minimum promoter (150b) and expressed canine FVIII in hemophilia A mice. Previous reports suggested AAV1 serotype is suitable for transduction of skeletal muscles and AAV8 serotype is superior to other AAV serotypes for transduction of the liver, thus, AAV1 vectors carrying the BDD cFVIII gene (AAV1 cFVIII) were injected to the skeletal muscles of hemophilia A mice. Since the liver could be transduced with intravenously injected AAV8 vectors carrying the Lac Z gene as efficiently as intraportally injected vectors, AAV8 vectors carrying the BDD cFVIII gene (AAV8 cFVIII) were injected to hemophilia A mice intravenously. FVIII clotting activities measured by the APTT method increased in mice injected with AAV1cFVIII in a dose dependent manner. The FVIII activity level in peripheral blood increased to 2.9±1.0% in hemophilia A mice with the AAV1cFVIII dose at 1x1012 gc/body, suggesting partial correction of phenotype with AAV1cFVIII vectors. The FVIII clotting activity levels in hemophilia A mice injected with AAV8cFVIII increased also in a dose dependent manner, achieving supernormal FVIII levels (436±219 %) in hemophilia A mice with the AAV8 cFVIII dose at 1x1012 gc/body. It is apparent that transduction of the liver with AAV8cFVIII is superior to transduction of skeletal muscles with AAV1cFVIII regarding FVIII production, however, AAV1 vectors have advantages of removing the transgenes in case of inconvenience and of less systemic distribution of vectors. These data suggested that both AAV1 and AAV8 vectors carrying the FVIII gene utilizing a minimum promoter have a potential for hemophilia A gene therapy.
Molecular Mechanism of Hepcidin Deficiency in a Patient with Juvenile Hemochromatosis.
Authors: Rideau Alexandra , Mangeat Bastien , Matthes Thomas , Trono Didier , Beris Photis 
Published: October 2019

Abstract
In eukaryotes protein translation is initiated at an AUG codon typically located within a so-called Kozak sequence (GCCRCC AUG G), a motif that presumably promotes the correct positioning of the mRNA in the ribosome. Recently, we described a patient suffering from juvenile hemochromatosis resulting from a lack of hepcidin production (Blood;15 June 2004; Epub ahead of print). We further revealed that this defect coincided with a G-to-A point mutation at position +14 of the 5′untranslated region (5′UTR) of the HAMP gene. This nucleotide change led to the creation of an ATG codon, out of frame with the physiological pre-hepcidin initiator located at positions +39-41. We speculated that this new ATG, which was embedded within a Kozak consensus sequence, acted as an aberrant translation initiation site that precluded hepcidin synthesis. In order to verify this hypothesis, we cloned in a mammalian expression vector the wild-type and mutant HAMP 5′UTR upstream of a modified EGFP cDNA, replacing the EGFP ATG by the HAMP physiological initiation codon. The resulting vectors were transfected into 293T cells. Presence of the additional ATG in the HAMP 5′UTR led to a 93 % decrease in GFP expression compared with wild-type. To confirm that this effect resulted from aberrant translation initiation at this site, a one-nucleotide deletion was introduced at the very 5′ end of the EGFP coding sequence, in order to place it in frame with the mutant HAMP 5′UTR proximal ATG (GTG in the wild-type sequence). This restored GFP expression to levels obtained when its coding sequence was in frame with the physiological hepcidin ATG of a wild-type HAMP 5′UTR. Our results very strongly suggest that the G to A mutation at position +14 of the HAMP gene 5′UTR blocks hepcidin production by inducing aberrant translational initiation of the pre-hepcidin mRNA. Whether a stable protein is synthesized from the mutated HAMP mRNA, which does not contain a stop codon in frame with the +14 AUG, remains to be determined. It is however unlikely that such protein contributes to the pathology observed in our patient, since the two other cases of hepcidin deficiency reported so far exhibit the same phenotype as our patient in spite of harbouring different mutations in the HAMP gene.
Hemojuvelin Mutations in Whites, Blacks and Asians with Primary Iron Overload and in Control Subjects.
Authors: Lee Pauline L. , Barton James C. , Brandhagen David J. , Niyongere Sandrine , Bohannon Sean , Rivers Charles A. , Acton Ronald A. , Beutler Ernest 
Published: October 2019

Abstract
Mutations in the hemojuvelin gene (HJV) are a cause of juvenile hemochromatosis (JH), but do they influence the phenotype of patients with adult-onset hemochromatosis with or without mutations of the HFE gene? We sequenced the coding region of the HJV gene of 133 subjects with primary iron overload (75 whites: 50 HFE, 25 non-HFE associated; 51 blacks: 1 HFE-associated; 7 Asians). A 63 year old white woman with non-HFE iron overload was found to be a compound heterozygote for HJV mutations: one previously identified (G320V); the other novel (C321W). The patient was diagnosed at age 30 (SI 223 μg/100 mL, TIBC 293 μg/100 mL, TS 76%, hepatic iron 31,154 μg/g, hepatic iron index 19.2). She had 2 healthy children, developed amenorrhea at age 23, had a history of hypothyroidism and developed Type 2 diabetes at age 59. She was iron depleted after 66 phlebotomies. Among 74 other white subjects with iron overload we found 3 polymorphisms: IVS2-6 C>G (3 subjects); nt 165 T>A (G55G), and IVS4-50 C>T (1 subject, each). Among 51 black iron overloaded and 132 black control subjects, the most notable polymorphism was a DNA triplet insert predicting an insertion of glycine. This was found in 2 subjects, one with and one without iron storage disease. Among the iron overloaded black subjects, we also found IVS2-6 C>G (1 subject), nt -140 G>C (3 subjects), nt 792 G>C (S264S) (1 subject), nt 828 C>A (I275I) (1 subject), and nt 929 C>G (A310G) (2 subjects). These polymorphisms, excluding S264S and I275I, were found in control subjects as well. No coding region mutations or polymorphisms were found in the 7 Asian iron overloaded subjects. We also determined the gene frequency of the HJV I222N and G320V mutations in adult control subjects from central Alabama, where a JH index case had been found with these mutations. Allele frequencies of I222N and G320V were 0.002 (n=240) and 0 (n=241) respectively in control whites. A white control female heterozygous for the I222N was also heterozygous for HFE C282Y but had normal iron measures. No I222N and G320V mutations were found in 124 and 118 black subjects respectively. We conclude that: 1) iron overloaded patients without HFE mutations, with or without a JH phenotype should be screened for HJV; 2) HJV mutations do not account for most cases of increased penetrance of HFE-associated hemochromatosis; 3) double heterozygosity for HFE C282Y and HJV I222N does not necessarily cause iron overload.
Geographic and Racial/Ethnic Differences in HFE Mutation Frequencies and Iron Phenotypes in the Hemochromatosis and Iron Overload Screening (HEIRS) Study.
Authors: Acton Ronald T. , Snively Beverly M. , Barton James C. , Adams Paul C. , Eckfeldt John H. , Harris Emily L. , Dawkins Fitzroy W. , McLaren Christine E. , Gordeuk Victor R. , McLaren Gordon D. , Reboussin David M. , Sholinsky Phyliss , Speechley Mark R. , Holup Joan , Leiendecker-Foster Cathy 
Published: October 2019

Abstract
Designing an optimal screening approach for hemochromatosis and iron overload requires knowledge of racial/ethnic subpopulations and frequencies of HFE mutations and phenotypes in geographic areas. In the HEIRS Study, HFE C282Y and H63D genotypes and prevalences of participants who met biochemical criteria for further evaluation (transferrin saturation >50% and ferritin >300 ng/mL, men; >45% and >200 ng/mL, women) were compared in a primary care-based sample of ~100,000 adults ≥25 years from 5 Field Centers. There were different respective HFE C282Y and H63D genotype frequencies in Whites, Blacks, Asians, and Hispanics across geographic areas (all p<0.0001), but not in Native Americans or Pacific Islanders. In Whites, the C282Y/C282Y proportion was significantly higher in AL than in CA or OR/HI. AL also had a significantly higher proportion of C282Y/wt than CA or OR/HI. In Blacks, D.C. had a significantly higher proportion of C282Y and H63D genotypes than AL. ONT had a significantly higher H63D genotype proportion than AL. In Asians, CA had a significantly higher proportion of H63D genotypes than OR/HI or ONT. In Hispanics, there were significant differences between: AL and CA; AL and D.C.; CA and D.C.; CA and OR/HI; D.C. and OR/HI; and D.C. and ONT. We also observed:
Prevalences of Participants Who Met Biochemical Criteria for Further Evaluation
Race/Ethnicity Geographic Area Prevalence (%) 95% CI Asians California 5.54 4.98, 6.16 Oregon/Hawaii 5.06 4.29, 5.95 Ontario 4.25 3.69, 4.88 Alabama 1.41 0.25, 7.56 D.C. 0.96 0.26, 3.42 Pacific Islanders California 3.66 1.25, 10.21 Oregon/Hawaii 3.47 2.23, 5.36 Native Americans Alabama 2.65 0.91, 7.52 Oregon/Hawaii 2.50 0.69, 8.66 D.C. 0.99 0.17, 5.40 Ontario 0.75 0.21, 2.71 Whites Ontario 2.31 2.08, 2.56 California 1.93 1.56, 2.37 Oregon/Hawaii 1.52 1.34, 1.73 Alabama 1.48 1.26, 1.74 D.C. 1.40 0.80, 2.43 Hispanics Ontario 1.92 0.82, 4.41 California 1.52 1.28, 1.79 Oregon/Hawaii 1.34 0.73, 2.4 Alabama 0.84 0.23, 3.01 D.C. 0.74 0.48, 1.16 Blacks D.C. 1.75 1.56, 1.96 Ontario 1.55 0.52, 4.45 Alabama 1.09 0.90, 1.32 Oregon/Hawaii 0.92 0.31, 2.66 California 0.86 0.29, 2.51
The percentage of men who met biochemical criteria for further evaluation was greater than that of women. We conclude that 1) C282Y and H63D genotype frequencies vary significantly within a single racial/ethnic group across geographic regions; and 2) the percentage of participants who met biochemical criteria for further evaluation varies by racial/ethnic group.