bluuebunny/crossref_metadata_2025 huggingface dataset:
Polymorphism in the Promoter Region of the Hemeoxygenase I Gene of the Donor Influences Overall Survival and Graft Versus Host Disease.
Authors: Gerbitz Armin , Hillemanns Patrick , Schmid Christoph , Wilke Andrea , Jayaraman Rajshri , Kolb Hans-Jochem , Bremm Heike , Eissner Guenther , Holler Ernst
Published: October 2019

Genetic polymorphisms of various donor and host genes have been found to be associated risk factors for graft versus host disease and transplant related mortality. Most of these genes are involved in the regulation of immune response and inflammatory reactions towards pathogens. The heme oxygenase (HO) protein is the rate limiting enzyme in heme degradation to biliverdin, free iron and carbon monoxide. Its inducible form I (HO-I) is constitutively expressed in the spleen due to the abundance of its substrate. It is inducible in a wide range of tissues upon exposure to cellular stress factors such as irradiation, bacterial lipopolysaccharide, proinflammatory cytokines and heat shock. Further, HO-I is involved in regulating inflammatory response and has been described as a “protective gene” in solid organ transplantation. In man the promoter region of HO-I displays length polymorphism due to a variable number of gt repeats within a 500bp region upstream of the TATA box. Individuals exhibiting 25 or less gt repeats express HO-I upon cellular stress at a higher level than individuals with more than 30 gt repeats. We retrospectively analysed length polymorphisms of 92 donor and host pairs undergoing allogeneic stem cell transplantation for various malignancies. We show here that donor promoter length polymorphism leading to low expression of HO-1 (>30 gt repeats) is associated with improved overall survival, relative to donors with ≤ 30 repeats (80.6% versus 54.4%, p=0.023). When subgrouped, having HLA identical siblings as donors had no influence on outcome (see figure below, right panel) whereas having matched unrelated donors strongly influenced overall survival (82.6 vs. 44.2%, p=0.015). Combination of high HO-1 expressing unrelated donors and high expressing hosts displayed a 27.8% survival (left panel, solid line vs.81.3% for individuals with one allele >30 repeats dotted line). Polymorphisms of the recipient HO-1 genes did not influence outcome after transplantation. When matched unrelated donors were used, the incidence of grade III gut GvHD was significantly higher when (p=0.032) and overall GvHD (≥ grade III) displayed a trend towards significance (p=0.098). Transplant related mortality was strongly influenced by having a donor who expresses HO-1 at a high level upon cellular stress (p=0.006). In a multivariate Cox regression analysis of pretransplant factors such as age, disease stage, sibling/unrelated donor etc. polymorphism in the donor HO-1 gene had a significant influence on overall survival (p=0.027). These data suggest a role for HO-I in GvHD pathophysiology and a possible role for HO-1 as a non MHC risk factor for unrelated transplantation.

Figure Figure
Serum Proteomic Profiling and Study of Haptoglobin Polymorphisms in Patients with Chronic GVHD after Allogeneic Hematopoietic Stem Cell Transplantation.
Authors: McGuirk Joseph P. , Yan Ying , Gang Hao , Hou Weijian , Abhyankar Sunil , Williams Casey , Belt Robert , Ruan Jianpa , Yan Weisi , Wieman Jeffery
Published: October 2019

We studied serum proteomic profiling in an attempt to discover novel biomarkers in the setting of clinically manifest graft versus host disease (GVHD) after allogeneic hematopoietic cell transplantation (Allo-HCT) by two-dimension gel electrophoresis (2-DE) and mass spectrometry analysis. More than 1000 protein spots per gel could be detected in individual samples and the distribution patterns of these spots differed significantly between the samples collected pre- and post- chronic GVHD (cGVHD) in the same patients. Mass spectrometric analysis in the paired 2DE gels from individual patients pre- and post-GVHD were performed. A group of proteins, Haptoglobin (Hp), Alpha-1-antitrypsin, Apolipoprotein A-IV, Serum paraoxonase and Zn-alpha-glycoprotein were demonstrated to be up-regulated and the proteins, Clusterin precursor, Alpha-2-macroglobulin, Serum amyloid protein precursor, Sex hormone-binding globulin, Serotransferrin and Complement C4 were found to be down-regulated in the patients with extensive cGVHD. Serum Hp was significantly increased in most patients following the development of cGVHD; Hp has been demonstrated to serve as an immuno-modulatory acute phase protein and may involve T- cell activation; further, Hp specific phenotypes have defined roles in influencing the pathophysiology of several immune system disorders. For these reasons, we used immunoblotting and PCR in combination with 2-DE gel image analysis to determine Hp polymorphisms in 24 allo-HCT patients (14 chronic GVHD, 4 acute GVHD and 6 without GVHD) and 10 normal donors. The results demonstrate that patients with cGVHD had a higher incidence of HP 2-2 phenotype (50.0%), in comparison to the patients without cGVHD (0%) and normal donors (10%) (Table 1), suggesting that the specific Hp polymorphism may play a role in the development of cGVHD after allo-HCT. A prospective analysis of Hp phenotype and the subsequent development of cGVHD is now being planned. This limited analysis suggests that proteomic profiling may provide significant contributions not only for identification of novel prognostic biomarkers, but also for novel insights into the pathophysiologic mechanisms of GVHD in allo-HCT.

HP Phenotype Patients 1-1 2-1 2-2 Chronic GVHD n=14 2(14.3%) 5(35.7%) 7(50.0%) Acute GVHD n=4 0 3 1 no GVHD n=6 1(16.7%) 5 (83.3%) 0 Normal Donor n=10 1(10.0%) 8(80.0%) 1(10.0%)
Impact of gamma/delta TCR+, Foxp3+ and Toll-Like Receptor Expressing T-Cells on the Outcome of Patients Receiving Unrelated Peripheral Blood Stem Cell Transplantation.
Authors: Platzbecker Uwe , Pabst Caroline , Schirutschke Holger , Wagner Ines , Theuser Cathrin , Ehninger Gerhard , Bornhauser Martin
Published: October 2019

Introduction: The role of gamma/delta (GD), foxp3 (regulatory, TREGs) and Toll-like receptor (TLR) expressing T-cells contained in the graft for the induction of graft versus host disease (GVHD) in patients after unrelated peripheral blood stem cell transplantation (PBSCT) is unknown.

Methods and patients: We therefore determined by flowcytometry and PCR the content of these T-cell subsets within the graft of 63 patients with hematological malignancies receiving unrelated PBSCT followed by cyclosporine A and methotrexate as GVHD prophylaxis and correlated these data with clinical endpoints. Patient and donor pairs were matched at least in nine out of ten characteristics (n=17), whereas a complete match (10 out of 10) was observed in 46 (73%) patients.

Results: The grafts contained a median of 11.25 x 106/kg (2.7–33.5) CD4+foxp3 T-cells and 9.79 x 106/kg (1.4–36.2) GD T-cells. Patients receiving lower amounts of CD4+foxp3+ cells had an increased risk of acute GVHD II–IV (65% vs. 44%, p=0.03). Whereas the overall CD3+ cell dose was not associated with the incidence of GVHD, increased counts of gdTCR expressing CD3+ cells showed significant correlation with an increased risk of GVHD II–IV (66% vs. 40%, p=0.028). A lower CD34+ cell dose was also significantly associated with an increased risk of GVHD II–IV (63% vs. 38%, p=0.04). Interestingly, there was a positive correlation between CD34+ and CD4+foxp3 cell dose (r=0.34, p=0.007).

In contrast to the blood of healthy donors, TLR2 and 4 were both expressed on up to 11 % of CD4+ and CD8+ cells within the graft but had no influence on the severity of GVHD. Interestingly, a higher expression of TLR2 and 4 on CD4+ but not CD8+ T-cells was associated with a higher treatment-related mortality (CD4: 17% for TLR2low vs. 45% for TLR2high, 13% for TLR4low vs. 52% for TLR4high, p=0.002).

Conclusion: Strategies aiming at selective manipulation of graft composition or add-back of distinct T-cell subsets after unrelated PBSCT might be a promising strategy in order to modulate the extent of clinical GVHD. Furthermore, up-regulation of TLR2 and 4 on T-cells and their role for non-relapse mortality warrants further investigation.
Translational Regulation of Gene Expression by the NPM-ALK Fusion Kinase of Anaplastic Large Cell Lymphoma: A Global Evaluation of Differentially Recruited mRNA’s to Polyribosomes.
Authors: Funato Kotaro , Haq Naznin , Sun Yezhou , Chen Jin , Khitrov Greg , Zhang Weijia , Sternberg David W.
Published: October 2019

The NPM-ALK fusion tyrosine kinase generated by the t(2;5)(p23;q35) chromosomal translocation is present in approximately 50% of anaplastic large cell lymphomas. NPM-ALK is known to play a causal role in the generation of hematopoietic neoplasia, and it has multiple potential sites of tyrosine phosphorylation that can bind to signaling effector molecules. We found that expression of NPM-ALK regulates mediators of translational initiation, including mTOR, S6K1, and 4E-BP1. Moreover, other investigators have shown that expression of proteins that mediate the control of translational initiation can be rate-limiting in the pathogenesis of neoplasia. Collectively, these findings stimulated our hypothesis that NPM-ALK promotes neoplasia by altering the partitioning of specific mRNA’s to polysomes and thereby regulating their translation. To evaluate this hypothesis, we assessed the global profile of polysomal mRNA in cells that express the NPM-ALK fusion kinase.

In this study, we generated Ba/F3 murine hematopoietic cells that express kinase-active NPM-ALK or a kinase-deficient NPM-ALK mutant (K210A). Cells were maintained in the presence of the cytokine IL-3 to minimize changes secondary to cell cycle redistribution or early apoptosis. We used these cell lines to compare in a global manner the partitioning of mRNA species to polysomes. We isolated polyribosome preparations using conventional sucrose gradient fractionation, and from this purified polysomal material we extracted mRNA. Similarly, we prepared total mRNA from these cell lines. The global profile of gene expression for both total and polysomal RNA was assessed using Affymetrix mouse genome microarrays. The data generated in this effort were analyzed using the GeneChip robust multichip analysis (GCRMA) method. Significance Analysis of Microarrays (SAM) and Linear Models for Microarray Data (Limma) were employed to identify significantly differentially expressed genes. By culturing each cell line in the presence of IL-3, we found little difference in total cellular mRNA levels. However, we found that expression of kinase-active NPM-ALK markedly altered the partitioning of mRNA’s to the polysomal pool. We identified 286 genes whose mRNA’s exhibited altered polysomal partitioning, including 102 decreased in cells with kinase-active NPM-ALK and 184 with increased polysome association. These translationally regulated mRNA’s encode proteins associated with transcriptional control, post-translational modification of proteins, and hematopoietic neoplasia. Our findings support the assertion that NPM-ALK signaling regulates gene expression in part through translational control. We speculate that concerted dysregulation of transcription and translation by oncogenes might broadly mediate the pathogenesis of hematopoietic neoplasia.
The Effect of Donor CD4+CD25+ Regulatory T Cells on Hematopoietic and Immune Reconstitution, GVHD and Disease-Free Survival after Allogeneic Hematopoietic Stem Cell Transplantation.
Authors: Yang Kai , Liu Qifa , Fan Zhiping , Sun Jing , Yi Zhengshan
Published: October 2019

Objectives To explore the effect of donor CD4+CD25+ regulatory T cells (TReg) on hematopoietic reconstitution, immune reconstitutuion and GVHD in patients who received allogeneic hematopoietic stem cell transplantation (allo-HSCT).

Methods 30 patients were divided into high TReg cell group (TReg≥10.0×106 cells/kg) and low TReg cell group (TReg<10.0×106 cells/kg) according to the number of TReg cells in the grafts. 13 patients were in high TReg cell group and 17 in low TReg cell group. The patients details of the two groups are in the following table. Flow cytometry (FCM) was used to continuously detect the percentage of TReg cells in the graft, T lymphocyte subsets and TReg cells in the recipients’ peripheral blood at different time after allo-HSCT. The hematopoietic reconstitution, immune reconstitution, TReg cell reconstitution, the incidence of GVHD and the disease-free survival were observed in the two groups.

Results The reconstitution of WBC and plantelet of high TReg cell group and low TReg cell group were 8.62±2.293 and 8.88±2.713, 12.69±5.736 and 15.18±16.708 days, respectively, and differed little between the two groups (P value was 0.778, 0.613, respectively). The incidence of acute GVHD (aGVHD) in high TReg cell group (15.38%) was obviously lower than that in low TReg cell group (58.82%) (P=0.016). There was significantly negative correlation (r=−0.382, P=0.037) between the number of infused donor TReg cells and severity of aGVHD. The reconstitution of CD4+CD3+T, CD45RO+CD4+T cells at 15 days and CD3+T, CD4+CD3+T cells at 30 days in high TReg cell group were notably faster than those in low TReg cell group after allo-HSCT (P value was 0.039, 0.024, 0.014, 0.020, respectively). The reconstitution of TReg cells at 15 and 180 days were significantly faster than those in low TReg cell group after allo-HSCT (P value was 0.013, 0.005, respectively). The 2-year disease-free survival of high TReg cell group and low TReg cell group were (69.90±15.50)% and (85.60±9.50)%, respectively, and there were no significantly difference between the two groups (P=0.514).

Conclusion TReg cells could contribute to decreasing the incidence of aGVHD, promoting immune reconstitution and TReg cells reconstitution.
Cyclosporin A Kinetics in Children after Stem Cell Transplantation.
Authors: Willemze Annemieke J. , Cremers Serge C. , Schoemaker Rik C. , Lankester Arjan C. , den Hartigh Jan , Burggraaf Koos , Egeler Maarten , Vossen Jaak M.
Published: October 2019

Introduction and aim of study. Cyclosporin A (CsA) is commonly used as prophylaxis against Graft versus Host disease (GVHD) after allogeneic haematopoietic stem cell transplantation (SCT). Therapeutic drug monitoring is necessary because of its unpredictable absorption and narrow therapeutic window. Usually dose adjustments are based on trough levels, although these correlate poorly with CsA systemic exposure in adults. The aim of this study was to investigate intravenous and oral CsA pharmacokinetics in children after SCT and to develop a limited sampling strategy in order to determine CsA systemic exposure.

Methods. Pharmacokinetics was investigated in children, aged 1.8 to 16.1 yrs, 2 to 43 days after stem cell infusion (median: 25 days), after intravenous or oral administration of CsA in, respectively, 9 and 8 children. Parameter estimation was performed using nonlinear mixed effect modeling as implemented in the NONMEM program.

Results and conclusions. Pharmacokinetics were described adequately with a two-compartment model with lag time (population estimates: Cl=10.5 l/h; Vc=15.5 L; Vp=60.6L; t ½ absorption=1.01 h, Tlag=0.6 h). Combination of Cthrough and a Bayesian fitting procedure with the pharmacokinetic model can adequately estimate the systemic exposure to CsA (r2=0.90, Figure 1). The estimation of actual AUC improves when more concentration time-points are used. No relation between body weight and clearance was found (Figure 2). Based on these data, CsA should not be dosed per kilogram body weight. A regimen with a fixed dose for all, adjusted on through levels in combination with the described pharmacokinetic model, may be more appropriate. (open circles: oral administration; closed circles: intravenous administration)

Figure Figure Figure Figure
Treosulfan, Fludarabine, and Antithymocyte Globulin - A Low Toxicity Conditioning Regimen for Unrelated Donor - Hematopoietic Stem Cell Transplantation in Chronic Myeloid Leukemia.
Authors: Holowiecki Jerzy , Giebel Sebastian , Wojnar Jerzy , Markiewicz Miroslaw , Holowiecka-Goral Aleksandra , Wylezol Iwona , Krawczyk-Kulis Malgorzata , Wozniczka Krzysztof
Published: October 2019

Unrelated donor - hematopoietic stem cell transplantation (URD-HSCT) is the treatment of proved long-term efficacy for chronic myeloid leukemia (CML) patients not having an HLA-identical sibling. However, high procedure-related toxicity observed after oral busulfan- or TBI-based conditioning limits its applicability and deteriorates outcome [Radich, Blood2003, 102, 31–5]. This is of increasing importance in the presence of challanging options offered by tyrosine kinase inhibitors. Between 2003–2006 we introduced a new preparetive regimen consisting of Treosulfan (a soluble alkylyting agent) 14 g/m2/d on days -6, -5, -4, Fludarabine 30 mg/m2/d on days -6, -5, -4, -3, -2, and, anti-thymocyte globulin (ATG) at a total dose of 6 mg/kg.

Thirty patients (age 32, range 16–48 years) with CML in the 1st chronic phase (n=29) or in 2nd chronic phase (n=1) were included in the study. Median interval from diagnosis to alloHSCT equaled 1.0 (0.5–12.0) years. 63% of patients had previously been treated with Imatinib. The donors were selected based on high resolution typing for both HLA class I and II. 43% of donors were mismatched for a single HLA-C (n=9), HLA-DQB1 (n=3) or HLA-B locus (n=1). Bone marrow was used a source of stem cells in 19 patients, peripheral blood - in 11 cases. GVHD prophylaxis consisted of Cyclosporin A and short-course Methotrexate.

All patients engrafted with the median time to neutrophil recovery >0.5 G/L and PLT >50 G/L of 19 (10–30) days and 18 (12–29) days, respectively. Complete donor chimerism was achieved until day +100 in all but one patient. Grade 3–4 neutropenic infections occurred in 13% of patients. Grade 3–4 mucositis as well as hepatic toxicity including VOD were not observed. The incidence of grade II acute GVHD was 23%, whereas grade III-IV acute GVHD was not observed. The incidence of extensive chronic GVHD was 10%.

At 3 years the probability of the overall survival and hematological relapse-free survival equaled 82% (+/−7%). The cumulative incidence of non-relapse moratlity was 18% (+/−7%) (fungal infection n=3, bacterial infection n=1, EBV-LPD n=1). Four patients required donor lymphocyte infusion or additional interferon or imatinib treatment because of incomplete donor chimerism or molecular/cytogenetic relapse after initial response.

We conclude that treosulfan + fludarabine + ATG conditioning is associated with low organ toxicity, low incidence of severe GVHD and NRM. The regimen is feasible option for CML patients referred for URD-HSCT in tyrosine kinase inhibitors era.
Pulmonary Changes in Myeloma Patients Following High Dose Melphalan.
Authors: Ford Clyde , Jensen Robert E. , Asch Julie , Pearl James E. , Crapo Robert O. , Petersen Finn
Published: October 2019

Background: Eligibility for high dose chemotherapy and autologous stem cell rescue often includes adequate pulmonary function. While high dose melphalan has been noted to cause significant pulmonary toxicity in a small percentage of patients with adequate pulmonary function, there are little published data on risk factors and use in lung impaired patients. We retrospectively reviewed the charts of 50 patients with myeloma who received a fixed dose of melphalan (200 mg/m2) and who had pre-melphalan and at least one post-melphalan spirometry (FVC and FEV1) and diffusing capacity (DLco) measurement. Comparisons were made with Pearson correlations and one-way ANOVA. Patients receiving melphalan showed no significant differences in mean FVC (p=0.40), FEV1 (p=0.48) or hemoglobin corrected DLco (p=0.40) comparing baseline to 3-month and 1-year results with 1-way ANOVA. There was a consistent increase in mean FVC(L), FEV1(L) and DLco (ml CO/min/mmHg) from pre to 3 months post (3m) and to one-year post (1y): FVC: pre=3.89, 3m= 4.02, 1y=4.24; FEV1: pre=2.92, 3m=2.95, 1y=3.14; DLco: pre=24.3, 3m=25.1, 1y=26.5. However, individual changes in DLco showed a wide distribution. Therefore, we examined a number of potential risk factors for change in DLco at 3 months. No detrimental effects were discernable for age, gender, history of prior pulmonary disease including smoking, pre-melphalan lung function, the use of high dose cyclophosphamide in the mobilization regimen, amount of prior chemotherapy or chest radiation, disease stage or response, or renal function. CD34+ cell dose (p=0.017, r = −0.344) and immunoglobulin type (IgA worse than IgG) (p=0.029, r = 0.316) were correlated and were independent factors on regression analysis (p<0.0018, r2=0.36). We conclude that on average, high dose melphalan produces little pulmonary toxicity and may be considered for selected patients with impaired baseline pulmonary function. It may be of value to avoid higher CD34+ cell doses in patients with a low pre-melphalan DLco. More work should be directed at identifying the factors that contribute to pulmonary function compromise following high dose regimens.
Prognostic Factors in the UK LRF CLL4 Trial.
Authors: Oscier David Graham , Wade Rachel , Orchard Jenny , Davis Zadie , Best Giles , Morilla Alison , Brito-Bapapulle Vasantha , Matutes Estella , Richards Sue , Catovsky Daniel
Published: October 2019

The LRF CLL4 trial randomised 777 previously untreated patients with Binet stage progressive A, B or C disease between January 1999 and October 2004 to receive either Chlorambucil, Fludarabine or Fludarabine and Cyclophosphamide. Interphase FISH for deletions of chromosome 6q, 11q, 13q, 17p and trisomy 12, IgVH gene mutational status (98% cut off), CD38 (7% cut off) and ZAP70 (10% cut off) expression were measured at randomisation on 579, 523, 535 and 478 patients respectively. Leukemic cells from 39 patients utilised the VH3-21 gene of whom 33 had homologous CDR3’s. Among the biological markers, log rank analysis showed that >20% p53 loss, del 11q, unmutated VH genes, high CD38 and high ZAP 70 correlated with disease progression or death (Table 1) but not deletion of chromosome 6q, 13q and trisomy 12 (p=0.7, 0.3 and 0.2 respectively). There was no difference in PFS or response duration between the 52 patients with 5–20% p53 loss and the 494 patients with no p53 loss. Multivariate Cox regression analysis showed that >20% p53 loss (p<0.0001), unmutated IgVH genes (p=0.0001), deletion of 11q (p=0.02) and male gender (p=0.03) were independent risk factors for short PFS. The effects of stage and age were overridden by FISH abnormalities. High ZAP70 expression was only significant when VH gene mutation status was not included in the model. CD38 expression was only significant in univariate analysis.

Table 1 Variable Progression or Death/N Univariate p-value(log rank test) Gender Male 384/573 0.002 Female 111/204 17q(p53) No 131/546 <0.00005 Yes 21/33 IgVH Unmutated 105/203 <0.00005 Mutated 225/320 del 11q No 288/463 <0.00005 Yes 87/116 ZAP70 Negative 140/242 0.003 Positive 158/236 CD38 Negative 110/201 0.0001 Positive 227/334

Among the 320 unmutated cases there was no significant difference in PFS between those with 100% homology (227 cases) and those with 99% or 98% homology to the germline sequence (93 cases). Mutated VH3-21 cases were more likely to express ZAP70 than other mutated cases, p=0.004. Excluding patients with >20% p53 loss, patients using the VH3-21 gene had similar progression free survival (PFS) to those remaining patients with unmutated VH genes and an inferior PFS to those with mutated VH genes (2p=0.0001). The adverse prognostic significance of 11q deletions was not clearly evident in an interim analysis presented at ASH ‘05. Patients can now be divided into 3 risk groups (Table 2). This risk stratification provides the basis of evaluating differing treatment modalities for each risk group in subsequent clinical trials.

Table 2 Risk Group Definition Progression or Death/N Univariate p-value (log-rank test) 3 yr PFS Poor >20%p53 loss 28/33 0% Standard Unmutated VH or 11q deletion or VH3-21 208/292 <0.00001 24.7% Good Mutated VH(excl VH3–21) 79/161 55.0%
Allogeneic Stem Cell Transplantation Should Be Performed in First Complete Remission in Adults with Acute Lymphoblastic Leukemia; Strong Antileukemic Activity of Chronic Graft-Versus-Host Disease.
Authors: Lee Seok , Cho Byung-Shik , Kim Sung-Yong , Eom Ki-Seong , Kim Yoo-Jin , Min Chang-Ki , Kim Hee-Je , Cho Seok-Goo , Kim Dong-Wook , Lee Jong-Wook , Min Woo-Sung , Park Jong-Won , Kim Chun-Choo
Published: October 2019

Purpose: The role of allogeneic stem cell transplantation (SCT) for adult acute lymphoblastic leukemia (ALL) remains unclear because interpretation of transplantation outcome is complicated by the criteria used to select patients for transplantation and by the relatively small number of patients studied. Moreover, whether SCT from an unrelated donor could be a treatment option of equal value in a case lacking a compatible related donor remains controversial. The aim of the present study was to determine the graft-versus-leukemia (GVL) effect and risk factors affecting outcome of 218 adults with ALL who received allogeneic SCT during the last 10 years (1995 to 2004).

Patients and Methods: The study population was 218 consecutive adults receiving an allogeneic SCT from matched sibling (n=162) or unrelated (n=56; 40 matched, 16 mismatched) donors at the Catholic Hematopoietic Stem Cell Transplantation Center in Korea. Their median age was 30 years (range, 15–61 years). One hundred eighty-three (83.9%) patients had high-risk criteria, and of these, 69 (31.7%) had t(9;22)/BCR-ABL and 7 (3.2%) had t(4;11)/MLL-AF4. One hundred sixty-five patients (75.7%) were transplanted in first complete remission (CR1); 23 (10.5%) in CR2; and 30 (13.8%) were resistant to chemotherapy before transplantation. Most patients (n=206, 94.5%) received a preparative treatment of total body irradiation (TBI)-containing regimen (TBI/cyclophosphamide for CR1, TBI/cytarabine/melphalan for >CR1). Graft-versus-host disease (GVHD) prophylaxis was attempted by administering calcineurin inhibitor (cyclosporine for sibling, tacrolimus for unrelated) plus methotrexate.

Results: With a median follow-up of 52 months (range, 15+ to 130+ months) after SCT, the 5-year probability of disease-free survival (DFS) was 51.3%±3.5% for all patients; 62.4%±4.3% for patients in CR1; and 11.3%±4.4% for patients in >CR1 at transplantation. There was no difference in DFS for sibling and unrelated transplant patients in CR1 (65.2%±4.3% v 62.3%±8.0%). Multivariate Cox regression analysis showed that the most powerful predictive factor affecting relapse and DFS was disease status at transplantation (CR1 v >CR1, p<0.001). The presence of chronic GVHD was also found to be significantly associated with favorable outcome (p<0.001).

Conclusion: Our data in combination with recent studies suggest that matched related or unrelated allogeneic SCT should be performed in CR1 in adults with ALL. Further studies to develop treatment strategies to reduce leukemic cell burden and to enhance GVL effect are needed. The indications for allogeneic SCT also should be continuously evaluated.
Abnormal Expression of Human Mitoferrin (SLC25A37) Is Associated with a Variant of Erythropoietic Protoporphyria.
Authors: Shaw George C. , Langer Nathaniel B. , Wang Yongming , Li Liangtao , Kaplan Jerry , Bloomer Joseph R. , Paw Barry H.
Published: October 2019

Mitoferrin (mfrn, slc25a37) is the high-affinity transporter of iron into mitochondria of developing erythroblasts and is essential for normal erythropoiesis. Recently, we described the cloning and characterization of mfrn in the zebrafish frascati (frs) mutant, in which homozygous null mutation leads to profound hypochromic anemia and embryonic lethality (Shaw GC, et al. 2006 Nature 440:96–100). Here we characterize the human MFRN orthologue and implicate a specific disease process purportedly caused by defects in MFRN expression. We studied human MFRN expression by a) analysis of human-tissue northern blots, b) fluorescent immunohistochemistry and confocal microscopy for subcellular localization, c) genetic complementation in MRS3/4 deficient yeast strains, and d) microinjection and rescue efficiency of frs mutant zebrafish. Human MFRN, mapping to chromosome 8p21, shows 82% and 64% similarity at the amino acid level to orthologous mouse and zebrafish Mfrn proteins respectively, and 60% similarity to a paralogous MFRN2 (SLC25A28). MFRN mRNA is highly expressed in fetal and adult hematopoietic tissues, including fetal liver and bone marrow, similar to zebrafish and mice. The MFRN protein localizes to the mitochondria like other SLC25 solute carriers. Ectopic expression of human MFRN capably restores hemoglobinized cells in anemic frs embryos, programs increased 55Fe-labeled heme synthesis in K562 cells, and rescues MRS3/4 mutant yeast for growth on iron-deficient media, demonstrating conservation of MFRN function. We evaluated 6 unrelated probands with a variant of erythropoietic protoporphyria (EPP) who harbor no mutations in ferrochelatase, the genetic cause of classic EPP. Five of the 6 index cases had severe hepatic disease necessitating liver transplantation. Our analysis of all 6 patients revealed the presence of an aberrantly spliced MFRN transcript, which was absent in normal individuals and patients with classic EPP. This aberrant spliceoform results in insertion of a 477 bp fragment from intron 2 and leads to an early nonsense codon after amino acid 156, prematurely truncating the MFRN protein. In contrast to normal MFRN cDNA, the variant MFRN cDNA from EPP patients fails to complement MRS3/4 mutant yeasts in low-iron media, demonstrating its loss-of-function activity. Our data demonstrate conserved function of human MFRN in erythroid mitochondrial iron metabolism and an association between a loss-of-function MFRN transcript and a variant of erythropoietic protoporphyria.
Favourable Overall Survival with Myeloablative Allogeneic Stem Cell Transplantation for Follicular Lymphoma.
Authors: Kuruvilla John , Bao Qikun , Messner Erica , Gupta Vikas , Kiss Thomas L. , Lipton Jeffrey H. , Minden Mark D. , Messner Hans A.
Published: October 2019

Introduction

Allogeneic stem cell transplantation (Allo-SCT) remains an option for patients with follicular lymphoma (FL). Our program employs a strategy using allo-SCT for patients (pts) with high risk disease based on clinical characteristics. We performed a retrospective analysis to examine long-term disease control and treatment-related mortality.

Methods

41 pts with indolent FL (follicular small cleaved [FSC], follicular mixed [FM] or FL grade 1,2 by WHO criteria) underwent allo-SCT at our institution between Jan 1989 and Dec 2005. Patients were in a chemosensitive remission at the time of SCT. The conditioning regimen consisted of busulfan (1 mg/kg PO q6h X4 days between 1989–200 and 3.2 mg/kg IV daily X 4days subsequently) and cyclophosphamide 60 mg/kg X 2 days. Cyclophosphamide 60 mg/kg X 2 days and TBI 12 Gy was used for unrelated donor SCT. GVHD prophylaxis was with cyclosporine A and methotrexate.

Results

There were 21 males and 20 females. The median age at the time of transplant was 45 years (range 24 to 58). Histologic subtype was: unclassified indolent FL: 3, Grade 1 or FSC: 17, Grade 2 or FM: 21. The median number of prior chemotherapy regimens was 2 (range 1 – 6) and was unavailable in 5. Prior anthracycline: 37, prior purine analog: 9, prior platinum-based: 20, prior auto-SCT: 1, prior rituximab: 11. The median time from diagnosis to allo-SCT was 23 months (range 6 – 161). 38 pts received BuCy conditioning. 2 patients underwent RIC SCT. Graft source was: matched related (MRD) bone marrow (BM): 28, MRD peripheral blood stem cells (PBSC): 4, Mismatch related (MMRD) bone marrow (BM): 1, MMRD PBSC: 2, matched unrelated donor (MUD) BM: 1, MUD PBSC: 1, and syngeneic BM: 4. The five year overall survival is 77% (95% confidence intervals 73 – 91%) with a median follow-up of 48 months post-SCT. Treatment-related mortality was 5 of 41 pts (12%). Non-relapse mortality was seen in one patient (3%). One patient has relapsed at over 3 years post-SCT while all recipients of syngeneic SCT remain in remission.

Conclusions

These results demonstrate that in selected patients, a fully myeloablative allo-SCT utilizing Busulfan-Cyclophosphamide conditioning provides excellent overall survival and disease control with low TRM. Prospective comparisons of RICSCT with myeloablative SCT should be performed in order to better evaluate these strategies.
Il-10 Polymorphism and TGF-β Haplotypes Are Associated with Rejection of Graft in Children with Beta Thalassemia Major Undergoing Matched Related Bone Marrow Transplantation (BMT).
Authors: Sellathamby Shanmugaapriya , Tamouza Ryad , Lakshmi Kavitha M , Viswabandya Auro , George Biju , Mathews Vikram , Krishnamoorthy Rajagopal , Charron Dominique , Chandy Mammen , Srivastava Alok
Published: October 2019

Th2 cytokines, IL-10 and TGF-β have immunoregulatory functions that can affect allograft acceptance. We have evaluated the correlation between IL-10 (-1082, -819, -592) and TGF-β codon 10C>T and 25G>C polymorphisms {(haplotype: TT/GG, TC/GG, high production); (haplotype: TC/GC, CC/GG, TT/GC, intermediate production); (haplotype: CC/GC, CC/CC, TT/CC, TC/CC, low production)}and graft rejection in 129 patients with β thalassaemia major who underwent matched related alloBMT. Patients who expired before day 15 (n=9) and for whom DNA samples was unavailable (n=18) were excluded from this study. None of these patients had graft rejection. Of the 102 patients who were evaluated, 73 (72%) patients received conditioning with Bu (16 mg/kg) / Cy (200 mg/kg) with ALG while 29 (28%) patients received Bu (600 mg/m2) / Cy (200 mg/kg) without ALG. All patients received unmanipulated bone marrow and standard cyclosporine / short course methotrexate regimen as prophylaxis against GVHD. Patients who died before day 21 after transplant (n=2) without achieving ANC >500/mm3 were not evaluated for engraftment. 9 out of 102 patients rejected their graft. Apart from the immunogenetic markers, the effect of other risk factors for rejection like recipient and donor age, sex mismatch, conditioning regimen, cell dose and Lucarelli class were also analyzed. Of these, only increasing donor age was found to mildly increase the risk of rejection (RR=1.06; p=0.019; 95% CI-1.01–1.1). Donor TGF-β haplotypes associated with intermediate or low production and IL-10 polymorphism accounting for low production were found to very significantly increase the risk of graft rejection (Table). TGF-β and IL-10 are essential for both development and the effector function of CD4 regulatory T cells which are important for tolerance induction. Therefore, it is possible that genotypes associated with lower production of these cytokines might contribute to increased graft rejection. We have previously reported that rejection in children with β thalassemia major is influenced by pharmacokinetics of busulfan and cyclophosphamide and genes implicated in their metabolism (BMT 2005 36, 839–845). We conclude that rejection of graft in this condition appears to be a multifactorial event affected not only by pharmacogenetics of the drugs used for conditioning but also by various immunogenetic factors.

Risk factors Relative risk P-value 95% CI TGFβ haplotypes Low + intermediate 6.97 0.015 1.4 – 33.6 High Reference IL-10 -819 TT 5.34 0.018 1.3 – 21.4 -819 TC + CC Reference IL-10 -592 AA 5.34 0.018 1.3 – 21.4 -592 AC + CC Reference
Conditioning with 8 Gy Total Body Irradiation and Fludarabine for Allogeneic Hematopoietic Stem Cell Transplantation in Acute Myeloid Leukemia: Projected 4-Year Update.
Authors: Stelljes Matthias , Bornhaeuser Martin , Kroger Matthias , Beyer Joerg , Sauerland Maria C. , Heinecke Achim , Berning Bjorna , Scheffold Christian , Silling Gerda , Buchner Thomas , Neubauer Andreas , Fauser Axel Arthur , Ehninger Gerhard , Berdel Wolfgang E. , Kienast Joachim
Published: October 2019

Seventy-one patients with acute myeloid leukemia (AML), most of them (63/71) considered ineligible for conventional allogeneic hematopoietic stem cell transplantation (HSCT), were enrolled into a phase II study on reduced intensity myeloablative conditioning with fractionated 8 Gy total body irradiation (TBI) and fludarabine (120 mg/m2) (Blood. 2005 Nov 1;106(9):3314–21). Patients received mobilized peripheral blood stem cells (n=68) or bone marrow (n=3) from siblings (n=39) or unrelated donors (n=32). HLA-typing was performed for HLA-A, -B, -Cw (serological matching or intermediate resolution DNA typing), DRB1 and DQB1 (high resolution DNA typing). Three patients had unrelated donors with an allele mismatch in HLA DRB1 (2 with an additional mismatch in HLA Cw) and 7 patients were transplanted from unrelated donors with an antigen mismatch in HLA Cw. Thirty-six patients were transplanted in complete remission (CR) and 35 with untreated or refractory disease (non-CR). Median patient age was 51 years (range, 20–66). Sustained engraftment was attained in all evaluable patients. With a median follow-up of now 41.3 months (range, 20.4–70.4) in surviving patients, probabilities of overall survival for patients transplanted in CR and non-CR were 80% (95% CI, 66 to 94%) and 17% (95% CI, 5 – 29%) at 4 years, respectively. Relapse-free survival rates were 57% (95% CI, 39 – 75%) and 14% (95% CI, 2 – 26%). Of the 35 evaluable patients transplanted in CR, 10 patients suffered a relapse between days 68 and 868 after transplantation (cumulative incidence 29%). Five patients with late relapse (>1 year after transplantation) achieved a subsequent CR after conventional chemotherapy, blood stem cell boost and treatment with granulocyte-macrophage colony-stimulating factor, lasting 2000+, 1841+, 909+, 847+ and 480 days, respectively. Depending on donor type, relapse-free survival was similar in patients transplanted from unrelated or sibling donors. Overall survival in patients transplanted in complete remission from unrelated vs. sibling donors was 84% (95% CI, 73 – 95%) vs. 77% (95% CI, 68 – 86%). The cumulative incidence of non-relapse mortality (NRM) in CR patients was 11% at 4 years and beyond (3 patients deceased before day 100 and 1 patient 25 months after transplantation), but amounted to 37% at 4 years in non-CR patients. Nine of the 33 surviving patients (27%) have actually active chronic GvHD (5 limited and 4 extensive disease). This update confirms that allogeneic HSCT from related or unrelated donors with 8 Gy TBI/fludarabine conditioning is feasible with low NRM and preserved long-term antileukemic activity in AML patients in first or later CR.
Long Remission Are Possible after Non Myeloablative Transplant in Patients with Follicular Non-Hodgkin’s Lymphoma (NHL): Results of Two Prospectives Multicenter Trials.
Authors: Caballero María-Dolores , Martino Rodrigo , Mateos María-Victoria , Briones Javier , de la Serna Javier , Tomás Jose-Francisco , Arranz Reyes , Díez Jose-Luis , Rubio Vicente , Ribera Josep-María , Moraleda José-María , Sanz Guillermo , Urbano Alvaro , Sarrá José , Sampol Antonia , Sierra Jordi , Miguel J. F. San
Published: October 2019

Myeloablative transplant has been investigated in poor prognosis indolent lymphoma; although recurrence rate is low it is associated with high mortality; the use of non-myeloablative conditioning regimens could reduce TRM maintaining the GVH effect. Up to May 2006, 35 patients with follicular NHL received a Non Myeloablative related allogeneic according to two prospective multicenter trials; conditioning regimen consisted of Fludarabine 150 mg and Melphalan 70–140 mg. GVHD prophylaxis consisted of CSA plus short-course MTX. All patients received filgrastim-stimulated peripheral blood stem cells from a HLA related identical donor. Median age at transplant was 50 years (34–62) and 16 (46%) had received a previous autologous transplant. At transplant, 5 patients (14%) were in CR1 (after several lines of chemotherapy), 9 (25%) in >CR1, 12 (34%) in PR, 1 (3%) had stable disease (after 3 chemotherapy lines) and 8 (23%) progressive disease. All patients engrafted. Acute GVHD developed in patients 19 (54%) (17patients (48%) grade II-IV). Chronic GVHD developed in 18 out of 27 patients at risk (67%), being extensive in 11 (41%). Disease was evaluated at day +100 and at that moment 23 patients were in CR, (85%) 1 (4%) in PR, two (7%) had stable disease and 9 patients ( 26%) have died. With a median follow up of 60 months (range: 32–80 months), 20 patients (57%) are alive disease free, and 14 (43%) have died, 12 of them (37%) due to transplant toxicity and 2 patients (6%) due to progression. Overall Survival (OS) and Event Free Survival (EFS) are 57 and 54 % respectively. Analysing variables which influence on OS and EFS, patients 55 years have a OS significantly shorter than those <55 years old (22% vs 69%; p:0,007). Moreover, patients who develop chronic GVHD have an EFS significantly better than those which do not develop it (78 % vs 44%; HR: 3,9 (1,0–14,6); p: 0,03). In conclusion, our results demonstrates high efficacy of non-myeloablative transplant in follicular NHL with a very low relapse rate, indicating the important role of chronic GVHD in the control of the disease; however, mortality rate is still high, mainly in patients 55 years. Although follicular lymphoma is considered as an incurable disease, our results after a large follow-up suggest that allogeneic effect could change this concept.
A Retrospective Single Institute Analysis of 127 Lymphoma Patients Who Underwent Allogeneic Stem Cell Transplantation: Impact on Peripheral T-Cell Lymphoma (PTCL) Including ATLL.
Authors: Tanosaki Ryuji , Kim Sung-Won , Yamazaki Satoshi , Fukuda Takahiro , Mori Shin-ichiro , Heike Yuji , Tobinai Kensei , Takaue Yoichi
Published: October 2019

Background Clinical application of allogeneic stem cell transplantation (allo-SCT) for lymphoma patients (pts) has been hampered by its high mortality rate. In order to explore and establish safe and efficient procedures, reduced-intensity stem cell transplantation (RIST) has been incorporated in recent protocols for lymphoma. Here we retrospectively analyzed our single institute experiences of allo-SCT for lymphoma.

Results Between 1999 and 2005, 127 allo-SCTs, including 94 RIST (74%), were performed for lymphoma in our single institute. The number of each pathological classification and the one in which RIST was used, which is shown in each parenthesis, were PTCL (non-ATLL) 15 (RIST 13), ATLL 25 (18), NK/T 3 (2), FL 36 (29), MCL 6 (5), DLBCL 23 (17), LPL 1(1), HL 5 (5) and LBL 13 (4). Most patients were not eligible for autologous transplantation (auto-SCT) because of relapse after auto-SCT or chemo-resistant disease. Transplant-related mortality (TRM) were 16% in RIST and 45% in conventional stem cell transplant (CST) (p<0.001), and it was as high as 54% in aggressive lymphoma pts treated with CST (n=26). 3-year OS and PFS of FL, PTCL including 2 ALCL and 1 AILT, and ATLL were 80.6±6.6% and 80.6±6.6%, 46.7±12.9% and 60.0±12.6%, 46.5±10.3 and 57.1±10.6%, respectively, with the median follow-up of 737 days (range, -4–2628) in all these pts. Pts with other histology had poor outcomes and the numbers of pts surviving more than 1 year were HL 1 (20%), MCL 2 (33%), DLBCL 7 (30%), NK/T 1 (33%) and LBL 3 (23%). 5 pts with DLBCL were treated with auto-SCT/RIST tandem transplantation according to in-house protocol, and 4 out of 5 were alive in current CR with median follow-up of 749 days (range, 580–1038).

Discussion Since RIST was associated with lower TRM than CST, it could be applied safely in heavily treated pts with refractory lymphoma. Most pts with DLBCL, NK/T, and LBL relapsed or died of disease. However, pts with FL, PTCL and ATLL, even in chemo-refractory cases, had durable remissions after RIST. In refractory DLBCL, de-bulking of the tumor by prior auto-SCT might be promising.

Conclusions RIST could be performed comparatively safely. Notably, PTCL, both ATLL and non-ATLL, were newly highlighted as a good indication of RIST and should be further investigated.

Figure Figure
Frontline High Dose Sequential Chemotherapy with Rituximab (R-HDS) and Autologous Stem Cell Transplantation Induces High Rates of Complete Response and Prolongs Survival in Mantle Cell Lymphoma (MCL).
Authors: Cortelazzo Sergio , Magni Michele , Pintimalli Milena , Mian Michael , Rossi Andrea , Billio Atto , Marchesi Maddalena , Zanni Manuela , Tarella Corrado , Boccadoro Mario , Andreini Angelo , Benedetti Fabio , Pizzolo Giovanni , Barbui Tiziano , Gianni Alessandro M. , Rambaldi Alessandro
Published: October 2019

We showed recently that the dismal outcome of MCL might be improved in 28 young patients (pts) (<61 years) by an up-front Rituximab supplemented high-dose sequential chemotherapy (R-HDS) supported by stem cell autograft (ASCT) (A.M. Gianni et al, Blood, 2003). Following this encouraging experience we treated other 26 pts aged<61 years with standard R-HDS and 19 aged >60 years with an age-adapted R-HDS. We report the outcome of 54 young pts (group 1), including the first 28 cases and that of 19 elderly pts (group 2). The majority of both groups had an advanced stage and bone marrow infiltration, while in elderly pts prevail B symptoms and > 1 extranodal site. One third of cases had >2 adverse prognostic features according to IPI. After 2–3 cycles of either doxorubicin- or cisplatin-containing chemotherapy, group 1 received standard R-HDS including: HD-cyclophosphamide (CTX) 7 gr/sqm and HD-Ara-C (2 g/sqm every 12 hours for 6 days). Following HDS chemotherapy a conditioning program with HD-melphalan (180 mg/sqm) and/or HD-mitoxantrone plus melphalan (60 and 180 mg/sqm) with ASCT was planned. Rituximab (375 mg /sqm) was given for a total of 6 doses, twice after HD-CTX and HD-Ara-C, as in vivo purging before CD34+ cells harvest and twice after ASCT. Elderly patients received an age-adapted R-HDS: HD-CTX (3–4 gr/sqm) and HD-Ara-C (1–1.5 g/sqm every 12 hours for 3–5 days), followed by HD-melphalan (120 mg/sqm) and HD-mitoxantrone plus melphalan (40 and 120 mg/sqm). 35 pts (65%) in group 1 and 9 (47%) in group 2 completed the planned program and a median number of 7.6 and 6.8 x 10^6 cells CD34+/kg were transplanted. After ASCT the CR rate was 88% in young and 95% in elderly patients. One young pt (2%) died during treatment, 1 developed sMDS and 5 died of late toxicity including a case of lung carcinoma. Among elderly pts only one died tardily because of cardiac disease. With a median follow-up of 48 months (range 8–101) in group 1 and 25 months (range 9–68) in group 2, the 5-year estimated OS, EFS and DFS were 77%, 60% and 71%, in group 1 and 55%, 53% and not yet achieved in group 2. The Cox multivariate analysis failed to identify within potential prognostic markers factors predictive for OS and EFS. We conclude that R-HDS is an effective regimen for the induction of complete remissions in pts with newly diagnosed MCL. The manageable toxicity of the program in elderly pts proved that an age-adjusted R-HDS regimen can be safely applied to this age subgroup still producing long-term remissions.