bluuebunny/crossref_metadata_2025 huggingface dataset:
Splice Form Specific Signaling of the Hematopoietic Growth Factor Receptor c-Kit.
Authors: Pedersen Malin , Sundberg Christina , Ronnstrand Lars
Published: October 2019

c-Kit is a member of the receptor tyrosine kinase family class III. Upon stimulation with its ligand, stem cell factor, c-Kit becomes activated and phosphorylated on tyrosine residues. The receptor activation leads to cellular responses such as proliferation, differentiation, survival and migration. c-Kit is known to be involved in the pathogenesis of several forms of cancer, e.g. gastrointestinal stroma tumors, leukemias and small-cell lung cancer. c-Kit exists in different isoforms due to alternative mRNA splicing. We are studying one pair of isoforms which varies by the presence or absence of four amino acids GNNK, in the juxtamembrane region just outside the plasma membrane. They are denoted GNNK− and GNNK+, respectively, and have previously been shown by our group to display functional differences in the activation of signaling pathways and cellular responses in the fibroblast cell line NIH3T3. The GNNK− splice form binds and activates Src family kinases more efficiently than the GNNK+ splice form. This leads to stronger phosphorylation of ShcA, extracellular signal regulated kinase 1/2 (Erk) and Cbl compared to the GNNK+ form. In order to study the two GNNK spliceforms in a more physiological context, we have retrovirally transduced two hematopoietic cell lines, 32D and BaF/3, which we now use as our model system. We could demonstrate that SCF stimulation induced a kinetically faster and stronger tyrosine phosphorylation pattern in the GNNK− splice form than the c-Kit GNNK+ splice form. Moreover, the phosphorylation of Erk and Shc is stronger in the c-Kit GNNK− splice form. These results are in line with the results described for NIH3T3 fibroblasts and previously published by our group. We are investigating the SCF-induced proliferative response in the hematopoietic cell lines 32D and BaF/3. Having established these model cell lines, we now aim to use Affymetrix microarray technology to detect differences in gene expression between the two splice forms upon ligand stimulation for different periods of time. As these different splice forms show remarkable differences in their signaling capacity we expect to find significant differences, but also similarities, in gene induction/repression mediated by the c-Kit splice forms. The idea is to link these findings to the biological responses induced by the two splice forms. We will more thoroughly investigate these findings using different c-Kit mutants and a battery of inhibitors of signaling proteins of interest available in the lab. An increased knowledge of c-Kit signaling in general and of the two different c-Kit splice forms in particular will help to find rational treatment for the unfavorable effect of c-Kit in cancer.
Mesenchymal Stem Cells, Fibroblasts and Pericytes: Different Functional States of the Same Cell?.
Authors: Covas Dimas T. , Penepucci Rodrigo A. , Fontes Aparecida M. , Orellana Maristela , Prata Karen L. , Neder Luciano , Perez Luiz C. , Carrara Rita C. , Araujo Amelia G. , Zago Marco A.
Published: October 2019

Mesenchymal stem cells (MSC) are multipotent precursors present in the bone marrow (BM), capable of differentiating into osteoblasts, adipocytes, chondrocytes and myoblasts. In addition to BM, MSC can be obtained from a variety of adult and fetal tissues. Additionally pericytes, that form a continuous subendothelial network of the vascular bed, share many properties with MSC, whereas the name stromal cell or fibroblast is often used interchangeably with MSC. How similar are MSCs obtained from different human tissues and what is their relationship with fibroblasts and pericytes? To answer these questions we have examined the expression of a set of 30 genes in 18 different human cell lines cultured in vitro: 7 MSC cultures from different sources (3 adult tissues: bone marrow, saphena vein, adipose tissue, and 4 fetal tissues: artery, liver, umbilical artery and umbilical vein), 2 of fibroblasts, 2 MSC cultures differentiated in vitro into adipocytes or osteoblasts, and one each of pericytes from the human retina, of bone marrow, endothelial cells, liver, brain, skeletal muscle and heart. Genes were selected on the basis of previous results of SAGE for MSC, CD34+ cells and fibroblasts. Cluster analysis using the software Cluster 3.0 UPGMA (average linkage) showed that all the MSC lines formed a very close cluster with pericytes and fibroblasts, separated from other normal human cells. Changes of the expression of highly expressed MSC genes were modest during in vitro differentiation of MSCs into osteoblasts or adipocytes, although the differentiation was accompanied by the increased expression of tissue specific genes. In addition, all the MSC lines had similar immunophenotypic markers and capacity for in vitro differentiation. Similarity of the gene expression profiles of MSC and fibrobasts were further confirmed by clustering of the 1,000 top expressed tags of SAGE libraries for 21 normal human tissues. Despite the similarity, expression of genes related to angiogenesis, especially CXCL6, was higher in MSC from umbilical vein, adult saphena vein and in pericytes than in BM-derived MSC. In conclusion, MSC that can be obtained from a variety of adult and fetal tissues have very similar biological markers, differentiation potential and gene expression profiles. Comparison of these characteristics also shows that MSC, fibroblasts and pericytes are very closely related, representing probably different functional states of the same cell. Finally, differentiaton of MSC into osteoblasts or adipocytes is marked by the expression of tissue specific genes, without dramatical changes of the overall profile.
Dicentric(7;9)(p11;p11) in Acute Lymphoblastic Leukemia: A Clinical and Laboratory Study of 7 Cases.
Authors: Pan Jin-lan , Xue Yong-quan , Wu Ya-fang , Wang Yong , Shen Juan
Published: October 2019

The clinical and experimental features of 7 cases of acute lymphoblastic leukemia(ALL) with the dicentric chromosome dic(7;9) (p11;p11) were investigated. Five patients were male and two were female. Their median age was 32 years. Cytogenetic examination of bone marrow cells was performed by direct method or 24h culture method. R banding techique was used for karyotype analysis. Among them, dic(7;9) was found to be the sole abnormality in 2 cases, t(9;22) and other additional aberrations were detected besides dic(7;9) in 4 cases as well as dic(7;9) and other abnormalities without t(9;22) were found in one case. Hyperleukocytosis(>100×109/L) was found in 4 cases with dic(7;9) and t(9;22), whereas 3 cases with sole dic(7;9) or dic(7;9) and other abnormalities had a WBC of 1.0–100×109/L. Enlargement of liver, spleen and/or lymph nodes were found in 6 cases. Immunophenotyping analysis showed that 5/6 cases of dic(7;9) ALL were B lineage ALL. Dual-color FISH detected BCR/ABL rearrangment in 3/6 cases and confirmed that the centromere of the derivative chromosome was originated from both chromosomes 7 and 9. A reciprocal translocation between chromosomes 7 and 9 was proved by chromosome patinting technique using whole chromosome paint probes for both chromosomes 7 and 9. This study has increased the number of reported cases of dic(7;9) from 8 to 15. Thus, we consider that dic(7;9) was a rare, but recurrent chromosome abnormality in ALL. It may appear as a primary change but also as secondary one following t(9;22), and had unique clinical and laboratory features.
Delivery of Autologous Bone Marrow Precursor Cells into the Spinal Cord Via Lumbar Puncture Technique in Patients with Spinal Cord Injury.
Authors: Callera Fernando
Published: October 2019

Alternative and less invasive routes for delivering hematopoietic stem cells in animals models of spinal cord injury (SCI) were described and the advantages of the percutaneous lumbar puncture (LP) technique have been recently demonstrated. (Bakshi A et al. 2004, Lepore AC et al. 2005) This proposition has not been described in humans. We therefore tested it by examining whether autologous bone marrow precursor cells can be delivered into the spinal cord via LP in patients with SCI. Seven patients with SCI (four had paraplegia and three had quadriplegia) were enrolled. The median age was 24 years and the mean duration of injury was 3 years. Patients received G-CSF (10microg/Kg) for five consecutive days and 100mL of bone marrow cells (BMC) were aspirated from the posterior iliac crest on day 6. Patients provided written informed consent. Mononuclear cells were separated by a Ficoll-Hypaque gradient, washed and resuspended in PBS. Flow cytometric enumeration of CD34+ cells was performed as previously described. (Gratama JW et al. 2001) Transplantation was performed 4 hours after bone marrow aspiration. CSF samples were collected before and seven days after the transplantation. For each patient, 15x106 mononuclear cells were injected via LP; the median number of CD34+ cells was 2.5x105 (1.8–5.0x105). Cell viability before transplantation was superior to 90% in all samples. Patients had no adverse events. CSF examination was normal in both times. Bone marrow precursor cells are clinically attractive because they can be obtained in patients at bedside raising the possibility of an autologous model of cell therapy for SCI. However, the potential therapeutic effects of these cells for SCI are poorly understood. What is the optimal CD34+ cell dosage? Will multiple doses of CD34+ cells be more efficacious? Is CD34+ the most appropriate BMC subset? Will BMC subsets combinations be more efficacious? The follow-up of our patients will probably provide some information. The absence of cells in CSF samples obtained after seven days is also intriguing. Although high speculative, a possible explanation is that cells home toward the injured spinal cord. All these issues need to be resolved in carefully designed experiments. Finally, our study demonstrated the possibility of delivering autologous bone marrow precursor cells via LP in patients with SCI. This procedure is feasible, safe and well-tolerated in humans. The potential therapeutic effects of these cells for SCI remain to be elucidated.
Effect of Plasma Exchange on Acute Graft Versus Host Disease after Allogeneic Hematopoietic Stem Cell Transplantation.
Authors: Cai Zhen , Zhen Gaofeng , He Huang , Li Li , Xie Wanzuo , Shi Jimin , He Jingsong , Lin Maofang
Published: October 2019

Acute graft versus host disease (aGVHD) is one of the major complications after allogenic hematopoietic stem cell transplantation (allo-HSCT) with high morbidity and mortality. Myeloablative treatment and the application of immuno-suppressive drugs (such as CsA, FK506 and MMF) are the main methods of prevention and cure. As it can separate pathogen, toxin, antigen and antibody from the plasma, plasma exchange (PE) is widely used to prevent and treat autoimmune disease, the rejection of stereo-organ transplantation, acute intoxication, severe hepatitis and acute hepatic failure. But up to now, PE is still rarely used to treat aGVHD. Here we present our review of cases where PE was accepted as a treatment modality for refractory aGVHD at our center. From October 2001 to March 2004, eight patients underwent PE due to aGVHD after Allo-HSCT with little response to immuno-suppressive drugs. Four of them had initial diagnosis of chronic myeloid leukemia (CML), and the other four were diagnosed with acute myeloid leukemia M2a,M4,M5,M6 respectively. The age of patients was from 14 to 38 years old (median 31). The “BuCY2” scheme (MMF+MTX+CsA) was used to prevent GVHD. After transplantation, all of them developed II-IV degree aGVHD. Each of them underwent 1 or 2 times of PE, with a volume ranging from 2500ml to 4000ml for a single PE. The overall response rate is favorable (fever, tetter, bellyache, diarrhea and liver function were ameliorated with various degree), with 3 cases (37.5%) having overt response, which was especially seen in patients with intestinal tract symptoms. Also, the level of TNF-α in the serum of the patients after plasma exchange was lower than that of the patients before plasma exchange(P=0.018). These results suggested that intestinal tract plays an important role in magnifying illness and “cytokine storm”, and TNF-α is the pivotal cytokine in the Intestinal tract GVHD. And the non-immunity aspects of GVHD can be described as an inflammatory process. PE may block this kind of inflammation by decreasing the cytokines, especially the level of TNF-α in serum after PE. Thus, the combination of PE and immuno-suppressive drugs is a rather good therapeutic method to control the symptoms of refractory aGVHD after allo-HSCT.
Evaluation of a New Program for PBSC Collection with Fresenius COM.TEC Blood Cell Separator.
Authors: Del Fante Claudia , Perotti Cesare , Viarengo Gianluca , Bergamaschi Paola , Marchesi Andrea , Parisi Cristina , Salvanesci Laura
Published: October 2019

Introduction: At moment PBSC collections can be performed using semiautomated or automated cell separator devices. The collection with semiautomated methods implies an augmented working load for the dedicated personnel and is strongly influenced by the operator. On the contrary, the automated methods offer the advantages of a diminuished working load for the dedicated personnel and an high standardization of the collection procedure. Herein we report our experience on 60 PBSC collections employing the new automated COM.TEC Fresenius autoMNC program that provides the possibility to predict the total number of CD34+ cells collected basing on the CD34+ cell count (x μL) pre-leukapheresis (LKF) collection in peripheral blood.

Materials and Methods: 39 patients affected with various onchohematological diseases and10 healty donors were mobilized with chemotherapy + G-CSF or G-CSF alone, respectively, and subsequently underwent LKF collection for auto or allotransplant. According to our internal protocol 60 LKF collections were performed starting with a CD34+ cell count in peripheral blood at least of 20/μL. Net weight of the final LKF product and its CD34+ cell content were evaluated at the end of each PBSC collection procedure and then compared to the expected data calculated by the cell separator device. Moreover a post collection peripheral blood Plt count was evaluated for each patient/donor.

Results: The mean starting WBC count was 25.86x103/μL (range: 4–82.3), Plt count was 151.38x103/μL (20–395), CD34+ cells was 96.63/μL (20–332). The mean WBC and CD34+ cells in the LKF collection were 224.78x103/μL (20.71–425.3) and 565.45x106 (59.3–1609.3), respectively. The mean volume of the LKF collection was 237.28 ml (120–503). The mean estimated CD34+ cell content was 498.37x106 while the real mean CD34+ LKF cell content was 623.32x106. The mean CD34+ cell collection efficiency was 91% (66–126). Finally, the mean post procedure Plt count in patient/donor was 77.91x103/μL (12–164).

Conclusions: The automatized PBSC collection with the new program COM.TEC Fresenius autoMNC demonstrated a very high CD34+ cell collection efficiency. Moreover the possibility to predict the CD34+ cell yield permits an optimal management of the LKF collection, reducing the number of procedures per patient/donor. The difference observed between the mean estimated CD34+ cells and the real CD34+ cell content may be due to the intra-procedure stem cell mobilization phenomenon. Finally, this new automatized collection system demonstrated to limit the collection related thrombocytopenia either in patient or in donor.
Cellular Composition of Allogeneic and Autologous PBPC Grafts and Patient Engraftment.
Authors: Barbosa Isabel L. , Benevides Patricia A. , Sousa Claudia C. , Machado Lopes Sergio B. , Tavares Ana R. , Campilho Fernando , Campos Antonio , Vaz Carlos P. , Sousa Susana R. , Pimentel Pedro , Carvalhais Alzira
Published: October 2019

We studied the influence of graft populations on the hematological recovery of PBPC transplants. The data shown is a retrospective study of 36 patients (median age 41, range 16–67) who underwent allogeneic transplant from HLA-matched siblings and 29 patients (median age 25, range 20–68) who underwent autologous PBPC transplants between June 2002 and July 2005. The cell populations studied were: total nucleated cells (TNC), CD34+ cells, T cells (CD3+) and NK cells (CD3−/CD56+). The hematological engraftment was evaluated by the following parameters: time to absolute neutrophil count (ANC)>0.5x109/L, platelets>20x109/L, absolute lymphocyte count (ALC)>0.5x109/L and ALC at day +15. The median cell doses infused in patients are summarised in Table 1

Table 1- PBPC cell population infused per Kg of patient body weight TNC x 108 CD34+ cells x 106 T cells x 108 NK cells x 106 *median **range Allogeneic 12.8 * 7.5 3.2 36.8 3.6 – 30.7 ** 3.4 – 10.5 0.7 – 9.8 4.2 – 136.6 Autologous 12 3.26 3.5 29 4.1– 51.0 2.2 – 7.0 0.1– 0.4 7.4 – 351

Spearman correlation analysis showed for the allogeneic PBPC grafts, correlation between TNC and NK cells collected (p=0.002), whereas for the autologous grafts there was correlation between TNC and NK cells (p=0.000) and CD34+ cells (p=0.004) collected.

To asses the importance of cell dose infused in the hematologic engraftment of the two patient groups, neutrophil, platelet and lymphocyte recovery was determined (Table 2).

Table 2- Patient engraftment Day to ANC>0.5x109/L Day to platelets>0.2x109/L Day to ALC>0.5x109/L ALC day +15 cell/μl *median **range Allogeneic 11 * 12 11 715 8 – 15 ** 8 – 29 2 – 14 190 – 4670 Autologous 11 11 12 1178 7 – 13 8 – 18 10 – 21 230 – 4410

For both groups of patients, engraftment occurred within the expected time and ALC day +15 cell/μl was different (median 715 cell/μl and 1178 cell/μl for allogeneic and autotransplanted patients respectively).

For the allogeneic transplanted patients, Spearman correlation analysis showed that the dose of NK cells infused was significantly correlated with ALC at day +15 (p=0.021). There was also correlation between the time to platelets and ANC engraftment (p=0.008) and time to platelets engraftment and ALC at day +15 (p=0.046).

For the autologous patients there was no correlation between cell populations of the grafts and the hematologic engraftment. Our data provide evidence that in allogeneic transplants the NK dose influences early ALC recovery. However, it is important to continue this study to balance the importance of cell population doses both in allogeneic and autologous transplants.
Intravenous (i.v.) or Oral (os) Busulphan (BUS) in Children Prior to Autologous (Auto) or Allogeneic (Allo) Stem Cell Transplantation (SCT): Pharmacokinetics (PK), Toxicity and Compliance.
Authors: Prete Arcangelo , Bartoli Antonella , Castellini Claudia , Rondelli Roberto , Masetti Riccardo , Pession Andrea , Regazzi-Bonora Mario
Published: October 2019

Introduction: High dose BUS is an important component of pre-transplant conditioning regimen in children with advanced hematological malignancies or solid tumor undergoing allogeneic or autologous stem cell transplantation. Variable intra and inter systemic drug exposure as measured by area under curve (AUC), can occurs following oral administration and may be influenced by age, body weight, absorption variability and, particularly in infants, by difficulties to assumption.

Aims: to compare the PK, the toxicity and the compliance of iv versus oral BUS administration in children undergoing pre-transplant conditioning regimen.

Patients and methods: 12 patients (pts), median age 37 (7–177) mos, body weight 6–69 kg, affected by AML (3), ALL (4), NB (3), Thalassemia (2), undergoing to autoSCT (4) or alloSCT (8), received as preparative regimen oral BUS at the dose of 1 mg/kg every 6 hours for a total of 16 doses (6 pts, GR1), or 2 hours i.v. infusion of BUS at the dose of 0.8 mg/kg every 6 hours for a total of 16 doses (6 pts, GR2). Four out of 6 GR1 pts (7–30 mos) had a nasogastric radiolus to allow BUS assumption. PK studies were performed after the first dose of BUS on blood samples collected at 0, 1, 2, 4, 6 hours after first administration in GR1 and at 0, 2, 3, 4, 6 hours from the beginning of the first infusion in GR2. BUS plasma level was determined by high-performance liquid chromatography as described by Henner et al.

Results: GR were comparable as for age (p= 0.07) and body weight (p= 0.14). Comparison of PK median values (+SD) in GR1 versus GR2 didn’t highlighted statistically significant differences as for AUC (p= 0.20) and for T1/2 (p= 0.11). Peak concentration was higher in GR2 (p= 0.058). Bioavailability was 0.61. Variation factor (VF%) were constantly low in GR2. No pt showed acute or late treatment-related extra-medullary toxicity and all pts developed a rapid and sustained hematopoietic recovery.

Conclusions: systemic exposure to 0.8 mg/kg iv BUS versus 1 mg/kg oral BUS is overlapping. Low VF% highlight a smaller interpatient variability in exposure compared to oral BUS. Moreover allows to avoid assumption difficulties in younger pts.
Detection of Cytomegalovirus Specific Cytotoxic T Lymphocyte in Allogeneic Hematopoietic Stem Cell Transplant Recipients.
Authors: Sun Jing , Huang Tao , Yi Zhengshan , Liu Qifa , Xu Dan , Ye Changxiong , Meng Fanyi
Published: October 2019

T lymphocyte reconstitution of allogeneic hematopoietic stem cell transplantation (alloHSCT) recipients was limited in the counts of T lymphocyte subgroup or T receptor recombination exclusion circles (TRECs). The function of T lymphocyte reconstitution is more important. Cytomegalovirus (CMV)-specific CD8 cytotoxic T lymphocytes (CTL) have the predominant role in protection against CMV. HLA-peptide Pentamer (Pentamer) and enzyme-linked immunospot (ELISPOT) have allowed direct visualization of CMV specific CTL. Both assays are highly specific and sensitive and had been applied to other areas.

29 patients with ANLL (9 cases), ALL (4 cases), CML (14 cases) and NHL (2 cases) were eligible for study. All patients received alloHSCT with 19 sibling-donors and 10 unrelative-donors in follow-up median 25 months, median age is 29 (range 11–52) and male/female is 16/13. CMV-IgG serologic status: 30 pairs were donor negative /recipient negative and 2 pairs were donor positive /recipient negative. The blood samples were collected from alloHSCT recipients with HLA-A2 at pre-HSCT and at 3, 6, 9, 12 month after alloHSCT. Detection of CMV-specific CTL in CD8 using HLA-A2-pp65495–503 pentamer complexes was performed with the fluorescence activated cell-sorter flow cytometer. IFN-gamma ELISPOT assay was performed in 96 bottomed-well microplates in which added 4×105 MNC /well and incubated with CMV pp65 (10ug/ml). T lymphocyte subgroup reconstitution was detected at same time.

24/29 (82.8%) recipients suffered CMV antigenemia once or more times after alloHSCT. The median time of onset CMV antigenemia is at 78d (range 31–476d) after alloHSCT. The counts of HLA-A2-pp65-specific CTL were continuously increased in 12 month after alloHSCT. The counts of HLA-A2-pp65-specific CTL at 3 month after alloHSCT were lower than at pre-HSCT and 6 month (p=0.000), 9 month (p=0.022), 12 month (p=0.013) post-HSCT obviously. The numbers of spots within 6 month post-HSCT detected for CMV-specific CTL by IFN-gamma ELISPOT was lower than that after 6 month post-HSCT (p=0.004). It mean that CMV-specific CTL grew more quick from 3–6 month than after 6 month. But analysis the reconstitution of CMV-specific CTL in every recipients were showed increasing or decreasing individually, which affected only by aGVHD (r=0.725, p=0.042) and cGVHD (r=0.843, p=0.036).
Benefit of Early Switch to Liposomal Amphotericin B in Patients with Amphotericin B Deoxycholate-Induced Nephrotoxicity Versus Late Switch.
Authors: Isnard Françoise , Laporte Jean P. , Chevallier Patrice , Pigneux Arnaud , Tilleul Patrick , Lafuma Antoine , Mahi Lamine , Deray Gilbert
Published: October 2019

Amphotericin B deoxycholate (AmBd) nephrotoxicity increases morbidity and mortality however there are no clear guidelines when to switch to liposomal amphotericin B (LAB). We evaluated the benefit of Early Switch (ES) to LAB in patients with AmBd compared to Late Switch (LS).

In this prospective randomized multicenter trial, standard AmBd 1mg/kg/d is initiated in neutropenic patients with haematological malignancies as empirical antifungal therapy (V0). If serum creatinine (SCr) increased of 30% from baseline V0, they were randomized (V1) to LAB 3mg/kg/d (ES) or to pursue AmBd (no switch). In a second step, AmBd treated patients were switched to LAB (LS) if their serum creatinine doubled from V0 or reached a value ≥ 170 μmol/l up to the end of treatment (V2). The analysis included 29 patients [ES=14; no switch=15 with late switch for 9 of them]. Demographics and mean duration of AmBd treatment prior randomization were similar in both groups. Early switch allowed a statistically significant (p=0.04) decrease in serum creatinine (−3.1%) from V1 to V2 as compared with a mean increase of +16.2% in the LS + no switch groups and also demonstrated a significant (p=0.04) increase (+5.1%) in creatinine clearance (ml/min) compared to a decrease (−10.3%) in the LS + no switch groups. This randomised trial demonstrated that an early switch to LAB improves and preserves renal function compared with late switch. Significant reduction of drug-induced nephrotoxicity may impact positively clinical outcomes and provide patients with greater options for chemotherapy treatments.

Evolution of renal function Criteria (mean) ES LS + no switch p SCr at V0 (μmol/l) 67.2 67.3 NS SCr at V1 (μmol/l) 109.0 108.2 NS SCr at V2 (μmol/l) 104.5 123.1 0.08 Evolution of SCr V1 - V2 (%) − 3.1 + 16.2 0.04 Evolution of ClCr V1 - V2 (%) + 5.1 − 10.3 0.04
Results of Non Myeloablative Geno-Identical Transplants in Adults with AML: Influence of the Doses of Peripheral Stem Cells Infused (on Behalf of the Acute Leukemia Working Party of EBMT).
Authors: Gorin Norbert C. , Labopin Myriam , Frassoni Francesco , Rocha Vanderson
Published: October 2019

In adult patients with acute myelocytic leukaemia receiving a conventional myeloablative allogeneic stem cell transplantation, several studies have shown that the higher the doses of hemopoietic stem cells infused, the better the outcome, with a lower transplant related mortality (TRM), a lower relapse incidence (RI) and a better leukaemia free survival (LFS). We wondered whether this observation was transposable in the context of non myeloablative transplants with peripheral blood stem cells where graft versus leukaemia (GVL) is the only anti tumor mechanism generated by the procedure in an effort to control the disease. In this particular situation, one might postulate that increasing the doses of peripheral blood T cells infused in an effort to increase GVL might increase GVHD and TRM. Further, one might speculate on the existence of an optimal range of stem cell and T cell doses to infuse. From January 1998 to December 2003, 253 patients with de novo AML, who received a non myelo ablative transplant with peripheral blood from a genoidentical donor, were reported to the ALWP registry with the dose of cells infused available. The sex ratio of the patients was 56% male /female. Patient age was 55 years (18–72). 141 patients were transplanted in first remission (CR1), 47 in second remission (CR2) and 65 patients had refractory disease or were transplanted in relapse. The pretransplant reduced intensity regimens were variable but 91% included fludarabin and Total body irradiation (TBI) was used in 23% of the patients at a dose<5 Gy. The median dose infused was 9.09 x 108 nucleated cells/kg (1.23–24). The follow up was 17 months (2–67). The LFS of the total population at 2 years was 41± 4%. For patients transplanted in CR1, CR2 and more advanced disease, the LFS were 49 ± 6%, 50± 8% and 23± 6% and the RI 37±4%, 34±7% and 69±6% respectively. Patients transplanted in CR had a significantly lower RI (p< 10−4, RR= 0.43 (0.29–0.65)) and better LFS (p< 10−5, RR=0.49 (0.35–0.7)) than those transplanted with advanced disease. Patients receiving doses of peripheral blood nucleated cells above the median had a trend towards a lower RI (42±5% versus 47±4%; p=0.06, RR=0.67 (0.45–1.02)) and a significantly higher LFS (46±5% versus 37±5%; p=0.04, RR= 0.68 (0.48–0.98)). Higher doses of nucleated cells were associated to a higher incidence of chronic GVHD (59 ± 5% versus 41 ± 5%, p=0.01, RR= 1.75 (1.12–2.73)). Interestingly, the use of TBI was associated to a higher RI (51± 8% versus 43±4%; p=0.02, RR=1.8 (1.1–2.9)). There was no relationship with the doses expressed in CD34+ cells infused. We conclude that in the context of non myeloablative transplants for adult AML, infusing higher doses of PB nucleated cells is beneficial to the patients.
Radiotherapy Post Autologous Stem Cell Transplantation in Patients with Lymphoma Not Reached Complete Response.
Authors: Chang Naibai , Gu Xichun , Zhu Ling , Wei Jianping , Zhao Shengming , Jin Liqing
Published: October 2019

Objective: Anthracycline-based chemotherapy induces 50%–70% of CR in patients with lymphoma, but only 30%–40% of long-term disease-free survival. Salvage chemotherapy with autologous stem cell rescue is required in patients with aggressive disease or never achieve CR with conventional chemotherapy,but the relapsed rate is still high. The purpose of this study was to evaluate radiotherapy post autologous stem cell transplantation in such group of patients.

Methods: 15 patients who underwent autologous stem cell transplantation during 1992–1998 were enrolled in this study. Conditioning regimen was CBV (cyclophosphomide + carmustine + etoposide). Radiotherapy was started on day +50(31–90). All patients were followed up until January 2005. Kaplan-Mier survival analysis was made by using SPSS10.0 software.

Results: There were 14 patients with non-Hodgkin lymphoma and 1 with Hodgkin disease enrolled. Male:female=11:4. Median age was 40 (30–64). At least 6 cycles of induction chemotherapy were given before transplantation. There were 3 patients in progression disease, 1 in stable disease, and 11 in partial remission before transplantation. Three patients received total lymph node irradiation (TLI). Seven patients received TBI(200cGY)+involved field irradiation therapy(IFIT). Five were treated with IFIT. All patients acheaved complete response(including 1 CRu) after radiotherapy. Three patients relapsed. One patient treated with TBI+IFIT relapsed at 6 months later. Two patients treated with IFIT relapsed at 8 and 36 months later respectively. The mean disease-free survival and overall survival were 10.84(SD1.37,95%CI) years and 11.89(SD1.35,95%CI) years respectively. The estimatrd 10-year disease-free survival and Overall survival were both 73%. One patient developed AML at 86 month. Grade III–IV hematologic toxicity was seen in 2 patients.

Conclusions: Post ASCT radiotherapy is safe and tolerated. Relaps rate is low. Patients who received TLI or TBI+IFIT seem to have better outcome than that received IFIT only.
Clinical Research of Autologous Hematopoietic Stem Cell Transplantation for Hematological Malignancies and Solid Tumors.
Authors: Huang Zhen-qian , Zhang Dong-hua , Tan Huo , Zhou Cheng-zhi , Liu Dan , Zhang Huan-zhu , Zhang Jie-xia , Huang Wei , Liu Wen-li
Published: October 2019

Objective: To evaluate the therapeutic effect of autologous hematopoietic stem cell transplantation (AHSCT) on hematological malignancies and solid tumors.

Methods: 20 patients with median age of 33.4±11.3 (18–50) years received AHSCT, 7 of them were acute non-lymphoblastic leukemias (ANLL)(CR1 5, CR2 1, refractory/relapse 1), 2 were acute lymphoblastic leukemia (ALL)(CR1 2), 1 was chronic myelogenous leukemia (CML-CP2), 1 was chronic lymphoblastic leukemia(CLL-NR), 6 were malignant lymphoma (CR1 2, CR2 2, NR 2), 1 was multiple myeloma, 1 was breast cancer relapsed after resection 10 years and lung and bone metastases, 1 was small cell lung cancer. 2 or 3 of following agents: Cytarabine(Ara-C)3–4g/m2, Cyclophosphamide (CTX) 4–6g/m2, Etoposide (VP-16) 0.5–1.0g/m2, Semustine (me-CCNU) 300mg/m2, Melphala n(Mel) 140mg/m2, Thiotep a (TSPA) 600mg/m2, Carboplatin (CBP) 1.0g/m2, were combined as conditioning regimen in all patients. Among them 2 patients with ALL accepted additional total body irradiation (TBI).

Results: All the patients have reconstituted bone marrow hematopoiesis after transplantation. None of them had the transplantation-related mortality. Among 20 cases, 15 achieved disease free survival (DFS) follow-up 36.5(2–106) months.

Conclusion: AHSCT might represent an effective approach for the treatment of some patients with chemosensitive solid tumor who are complete remission or part remission. Without compatible donors, patients with leukemia and malignant lymphoma at CR1 stage could receive AHSCT to reduce relapse and increase disease-free survival. It is suggest that have a obvious survival benefit from AHSCT.
Use of Complementary and Alternative Medicine (CAM) by Patients with Myeloma or Lymphoma.
Authors: García-Nieto Antonia , Capote F.J. , Martín-Reina M.V. , Bailén A. , Fernández-Valle M.C. , Gil J.L.
Published: October 2019

The use of complementary and alternative medicine (CAM) is wide-spread among cancer patients. Few studies have investigated the use of CAM by patients with haematological cancer patients in Europe.

OBJECTIVE: To investigate the distribution and patterns of CAM use by patients with myeloma or lymphoma.

PATIENTS AND METHODS: 103 patients with lymphoma and 23 with myeloma (72 male and 54 female) they have responded a questionnaire on diverse aspects of the use of CAM.

RESULTS: Ninety-six patients (76.1%) recognize to have used or to use some type or product CAM: Matricaria chamomilla L. (75,3%), Tilia vulgaris (57.4%), green tea (19.8%), Valeriana officinalis L (17.4%), Royal Jelly (17.4%), soya (16.6%), Brewer’s Yeast (7.9%), wheat germ (6.3%), ginseng (6.3%) and aloe vera (6.3%); Cat’s Claw, Hypericum Perforatum (St. John’s Wort), echinacea, grape seed, milk thistle (Carduus marianum), graviola and marijuana (less of 5%). Forty-nine patients (38.8%) recognize to have practiced or to practice oration (23.8%), relaxation (11.9%), oils massages (9.5%), meditation (6.3%), imposition of hands (3.9%) and yoga (3.9%); music therapy, homeopathy, Shiatsu, acupuncture, therapies through of the movement and the dance, guided imagination, psychotherapy, chiropractic and consult with healer (less of 3%). Visualization, hypnosis, biofeedback, chromotherapy, reflexology, or therapies through humour have not talked about or through Internet. They recognize to look for a complement for the traditional medicine (29.6%), to fight the indirect effect (17.4%) or to have control on the own disease (16.6%) not being felt impassive (23.8%); to content the family (3.1%) or to think that its treatment will be faster (21.4%) is other reasons. In 3.9% they tried to replace the traditional medicine. Only the 30.1% “complementary and alternative medicines” knew the meaning the terms. The 62.6% did not know the qualification of the personnel who sells products or applies treatments to him MAC and 36.6% not informs its doctor that take CAM. Sixty-one percent consider that the public system would have to finance this type of practices.

COMMENTARIES: This study contributes data on the use of therapies MAC by patients with myeloma or lymphoma in Spain. The professionals we must make the effort necessary to improve our knowledge of the MAC and the formation of the patients; the MAC that demonstrate to their effectiveness and security would have to be able to integrate themselves to the care of the patients.
Health-Related Quality of Life (HRQL) in Children with Severe, Chronic Immune Thrombocytopenia (cITP) Treated with Rituximab.
Authors: Parsons Susan , Bennett Carolyn , Mahoney Donald , Olson Thomas , Sawaf Hadi , Moore Theodore , Loh Mignon , Buchanan George , Neufeld Ellis
Published: October 2019

Background: A subset of children with cITP is at increased risk for severe bleeding and functional limitations. We evaluated the HRQL of children with cITP as part of a multicenter, open label Phase I/II clinical trial of four weekly doses of Rituximab, evaluated for response at 12 weeks. Responders had a ≥ 50,000/mm3 platelet count over weeks 9–12 without other supportive therapy.

Methods: All participating parents were invited to complete the Child Health Ratings Inventories (CHRIs) at baseline and 12 wks. The CHRIs, a generic HRQL questionnaire, contains 20 items, forming four domains in the areas of physical, role, and emotional functioning, and school. Each item has a five-point Likert response scale. Domain scores range from 0–100, with higher scores connoting better functioning. Of the 36 study participants, 24 parents opted to participate in the HRQL evaluation; 22 of 24 (91%) completed both the baseline and 12-week assessments. The baseline clinical profile and 12-week Rituximab response rates of HRQL participants and non participants (n=12) were compared. Potential clinical covariates (e.g., duration of illness, worst bleeding score, platelet count, number of prior therapies) were examined in a univariate analysis (UVA) with baseline HRQL domain scores. Regression analyses (MVA) adjusting for age and gender were used to assess the effects of treatment response and clinical covariates on the mean change in scores between baseline and week 12.

Results: While HRQL participants had more previous treatments and lower platelet counts than nonparticipants, their clinical profile was otherwise similar; Rituximab response rates did not differ. Mean HRQL domain scores (standard deviation) at baseline were as follows: physical-52.1 (36.2); role-78.6 (23.7); emotional-63.4 (18.3); school-64.1 (25.8). Lower platelet count and greater number of prior therapies were significantly associated with worse baseline HRQL scores on UVA. By week 12, 36% (8/22) responded to Rituximab. Clinically meaningful improvements in HRQL scores (≥ 0.5 standard deviation improvement) were observed in responders; no change was detected in non-responders. In MVA, responder status contributed the predominate effect on the HRQL change scores, with the greatest improvement noted in emotional functioning (p=.002). No other clinical variables emerged in this limited sample size.

Conclusions: The pattern of parent-reported HRQL scores for these children with severe cITP mirrored scores from other severe, pediatric health conditions. Significant improvements in HRQL were observed among Rituximab responders over the 12-week period while non-responders had no change. Further overall characterization of HRQL in the cITP pediatric population is needed.
Role of Atypical p50:p50:Bcl-3 NFκlB Complex in Differentiation of Erythroid Cells.
Authors: Sawyer Stephen , Chen Jingchun
Published: October 2019

We recently reported that mouse and human primary erythroid progenitor cells and erythroid cell lines synthesize and respond to Tumor Necrosis Factor-alpha (TNF-α). The nuclear transcriptional control complex, NFκlB is central in signaling downstream from TNF-α; so we began to study the function of NFκlB in erythroid cells. We made three very interesting initial findings: 1) first we found that NFκlB binding to DNA increased very slowly in HCD57 erythroid cells treated with erythropoietin (EPO, the hormone required for red blood cell development). An inhibitory effect of adding a neutralizing antibody to TNF-α on EPO-stimulated NFκlB DNA suggested this increase in NFκlB was due to TNF-α rather than direct EPO signaling. 2) We also found that NFκlB binding to DNA increased 10-fold or greater during erythroid differentiation. We found greatly increased NFκlB DNA binding in HCD57 cells that differentiated due to over-expression of JunB, F-MEL induced by DMSO, or human UT7-EPO or murine HCD57 cells induced to differentiate with hemin. 3) Surprisingly, we found that the NFκlB DNA binding complex in mouse primary erythroid cells and the erythroid cells lines tested was almost exclusively composed of the atypical p50/p50/Bcl3 NFκlB rather than the canonical p65/p50 or the non-canonical p65/p52 NFκlB. When we begin to study the biological significance of this atypical NFκlB in EPO-mediated erythroid differentiation in vivo using genetic tools, we found marked deficiencies in the development of erythroid cells in either the nfkb1−/− mice (p50−/−) or the bcl3 −/− mice. The nfkb1−/− mice were mildly anemic. The number of red blood cells in the circulation of these mice was statistically lower than in control mice. The number of CFU-e was also reduced in nfkb1−/− mice. Using the Ter-119 and CD71 staining method, we noted that proerythroblasts and immature erythroid cells increased and mature erythroblasts decreased in either non-anemic bone marrow or anemic spleens of nfkb1−/− mice. Forward scatter of Ter-119+ cells also showed an increased size of the average immature erythroid cell in the bone a marrow of nfkb1 −/− mice, suggesting a block in differentiation and continued cell cycling of the immature erythroblasts. Similar erythroid defects were observed in the spleens of anemic bcl-3−/− mice. nfkb1−/− mice and bcl-3−/− mice are also apparently unable to produce new reticulocytes as effectively as wild type mice after induction of anemia. Our working hypothesis is without expression of either p50 or Bcl-3 NFκlB proteins, immature erythroid cells continue to proliferate and ineffectively differentiate. In summary, the atypical p50/p50/Bcl-3 NFκlB complex appears necessary for maximal differentiation of immature erythroid cells.
Fusion to Vp16 Converts MEIS1 into an Oncoprotein That Immortalizes Progenitors and Causes AML in the Absence of Coexpressed Hox Genes: Hoxa7 and Hoxa9 Induced Further Stem Cell Gene Transcription in Vp16MEIS1 Progenitors.
Authors: Wang Gang G. , Pasillas Martina P. , Kamps Mark P.
Published: October 2019

MEIS1 and Hoxa9 are homeobox transcription factors that promote self-renewal in hematopoietic progenitors. MEIS1 does not induce leukemia, but cooperates strongly with Hoxa9 to produce acute myeloid leukemia (AML). Previously, we demonstrated that Hoxa9 blocks differentiation of myeloid progenitors that do not express MEIS1 and do not induce leukemic. Coexpression of MEIS1 causes transcription of genes that segregate with the leukemia-initiating subset of human AML blasts, such as CD34 and FLT3. We designate these genes as leukemic stem cell genes, or LSC genes. MEIS1 promoted LCS gene transcription by a mechanism that requires interaction with PBX and DNA, and that also requires a short MEIS1 C-terminal transactivation domain (CTD). Here we use a dominant transactivating or transrepressing form of MEIS1 to determine whether the activation or repression function of Pbx:MEIS1 complexes is sufficient to cause myeloid leukemia in combination with coexpressed Hoxa9. Surprisingly, fusion of MEIS1 to the Vp16 transactivation domain (but not the engrailed transrepression domain) produced an autonomous oncoprotein that immortalized progenitors and caused myeloid leukemias without the need for coexpressed exogenous or endogenous Hox genes. Like MEIS1, Vp16MEIS1 required binding to Pbx and DNA for immortalization; however, the CTD was not necessary in the context of Vp16MEIS1. This suggests that the CTD participates in target gene activation in AML blasts, a function replaced by Vp16 in its absence. Retroviral expression of Hoxa9 or Hoxa7 induced a further, strong, transcriptional upregulation of LSC genes in Vp16MEIS1 progenitors and elevated their leukemic potential to the level of bona fide AML blasts. These data suggest that transactivation is the essential function of Pbx:MEIS1 complexes in AML, and that HOX proteins cooperate with Pbx:MEIS1 complexes to activate transcription of early progenitor genes whose expression is required for human AML.
Human CD4+ CD25high Regulatory T Cells Inhibit Myeloid but Not Plasmacytoid Dendritic Cells Activation.
Authors: Houot Roch , Perrot Ivan , Durand Isabelle , Garcia Eric , Lebecque Serge
Published: October 2019

CD4+CD25+ regulatory T cells (Treg) are essential negative regulators of immune responses. However, the mechanisms of immune suppression and the spectrum of cells they target remain incompletely defined. In particular, although Treg directly suppress conventional T cells in vitro, they might also affect antigen presenting cells (APC).

Here, we studied the maturation of human myeloid (mDC) and plasmacytoid (pDC) dendritic cells activated with Toll-like receptor (TLR) ligands in the presence of CD4+ CD25high regulatory T cells in vitro. T cells and DC subsets were purified from normal human peripheral blood. LPS, CpG ODN 2216 and R-848 were used to trigger the maturation of mDC, pDC or both through TLR4, TLR9 and TLR7/8 respectively. Preactivated CD4+ CD25high Treg had no effect on the maturation of pDC. Conversely, they strongly suppressed TLR-triggered mDC costimulatory molecules up-regulation, pro-inflammatory cytokines secretion and their antigen presentation capacity, as opposed to conventionnal T cells (Tconv). At a ratio of 3 Treg for 1 DC, the percentage of mDC acquiring CD80 was reduced 5 fold (from 75% to 16%) while the Mean Fluorescence Intensity was decreased by approximately 65% for CD80 and 35% for CD86 after LPS stimulation and by 50% and 20% after R-848 stimulation. Furthermore, Treg dramatically decreased the secretion of IL-12p40, TNF-α, and IL-6 by mDC (95%, 93% and 50% average inhibition respectively) after LPS activation and to a lesser but still significant extent (38%, 35%, and 38% average inhibition respectively) after R848 stimulation. Finally, we found that Treg-conditionned mDC had a reduced ability to trigger naïve T cell proliferation in a mixed leukocyte reaction.

Suppression of mDC activation by Treg appeared to require cell-cell contact. Moreover, the inhibition of pro-inflammatory cytokines secretion, but not of phenotypic maturation, was almost completely restored using an anti-IL10 receptor monoclonal antibody, but not anti-TGFβ nor anti-CTLA-4 blocking antibodies. Those data suggest that Treg prevent the co-stimulatory molecules up-regulation on mDC through contact dependent mechanisms, while the modulation of cytokines secretion appears to be largely mediated by IL-10.

Overall, our results provide the first evidence of a direct inhibition of human mDC but not pDC maturation by CD4+ CD25high Treg. Therefore, by restraining the maturation of mDC, human Treg may enlist those cells for the initiation and the amplification of tolerance in vivo.