bluuebunny/crossref_metadata_2025 huggingface dataset:
The Hematopoietic Cell Transplantation Comorbidity Index: No Effect on Outcome of Reduced-Intensity Allografts.
Authors: Frankfurt O. , Altman J. , Krishnamurthy J. , Evens Andrew M , Gordon L. , Singhal S. , Tallman M. , Williams S. , Winter J. , Duffey S. , Singh V. , Meagher R. , Mehta J.
Published: April 2020

The hematopoietic cell transplantation-comorbidity index (HCT-CI) has been proposed as a means to estimate the risk of transplant-related mortality (TRM) and the likelihood of overall survival (OS) after allogeneic HSCT. However, its value and applicability remain unclear; in a 2-center study, it was predictive of outcome at one but not at the other in multivariate analysis (Blood2007;110:4606).

We investigated the effect of HCT-CI in 130 adult patients with hematologic malignancies undergoing reduced-intensity HSCT after 100 mg/m2 melphalan (+ 50 mg/kg cyclophosphamide if no prior autograft). GVHD prophylaxis comprised cyclosporine/tacrolimus and MMF. 18% of the donor-patient pairs were 1 antigen/allele-mismatched. 57% had refractory disease. 45% had failed an autograft. ECOG performance status (PS) was 0–34%, 1–48%, 2–14%, and 3–4%. The HCT-CI score distribution was 0–10%, 1–4%, 2–28%, 3–24%, 4–15%, 5–10%, 6–8%, and ≥7–2%. Overall, the patient population had higher HCT-CI scores than described in other studies (Cancer2008;112:1992).

In multivariate analysis, TRM was higher with HLA mismatch and PS 2-3, and OS was lower with PS 2-3, HLA mismatch, refractory disease, donor age >45 y, elevated LDH, and platelets <100.

HCT-CI 0 vs 1-2 vs ≥3 had no effect on TRM or OS. HCT-CI 0-2 vs ≥3 had a borderline effect on TRM (P=0.09) and OS (P=0.15) in univariate analysis, but no effect in multivariate analysis when forced into the model with factors known to be significant. No significant correlation was seen between HCT-CI and any of the factors found to be predictive of outcome.

Figure Figure

Because serum albumin is a powerful non-specific predictor of general health and outcome in all clinical situations, we hypothesized that it would correlate with the PS. As shown in the figure below, it did. We expected that it would correlate with HCT-CI because increasing comorbidity should theoretically be associated with declining albumin. However, it did not.

Figure Figure

In our experience, HCT-CI does not correlate with outcome – especially when other pertinent factors are taken into account. Since most data supporting HCT-CI have originated from a single center, it is possible that its applicability depends on individual centers and the treatment protocols employed. However, lack of correlation of HCT-CI with albumin does call its biological plausibility into some question. HCT-CI cannot be employed unless refined and validated prospectively in multi-center studies.
Febrile Reactions Occurring with Second Cycle of High Dose Melphalan and Stem Cell Transplantation in Patients with AL Amyloidosis: A “Melphalan Recall” Reaction.
Authors: Rosenzweig Michael , Seldin David C. , Remick Daniel G , Skinner Martha , Quillen Karen , Oran Betul , Finn Kathleen T. , Sanchorawala Vaishali
Published: April 2020

Systemic AL amyloidosis is a clonal plasma cell disease related to multiple myeloma characterized by the wide spread deposition of amyloid fibrils into tissues and organs. Treatment for this disease is directed towards reducing amyloidogenic monoclonal light chains by attacking the underlying plasma cell dyscrasia. Aggressive treatment with high dose intravenous melphalan followed by autologous stem cell transplantation (HDM/SCT) is effective in inducing hematologic and clinical remissions and in extending survival. Tandem cycles of HDM/SCT have been shown to increase hematologic complete response (HCR) rates in patients with multiple myeloma and have therefore been included in the treatment of AL amyloidosis. Between April 1994 and July 2008, 57 patients with AL amyloidosis at Boston University Medical Center were treated with a second cycle of HDM/SCT after failing to achieve a HCR following a first transplantation. Eleven of 57 patients (19%) treated with tandem transplantation developed a high fever; 12-24 hours following melphalan administration. Other clinical features noted in some of the patients include hypotension, acute renal failure, and skin rash. Among the 11 patients described, there were 7 men and 4 women. The average age was 53.5 years (range 41-60). The average peak temperature of the 11 patients was 39.1º C (range, 38.1-39.4°C). All patients experienced resolution of fever within 24–48 hours. Six of the 11 patients developed a rash and 5 developed hypotension that was responsive to intravenous fluids or pressors when indicated. Two of the patients developed acute renal failure that improved following resolution of the fever. In all 11 patients, workup for an infectious etiology of fever was negative. One of the 11 patients described, had cytokine measurements before, during and after the febrile reaction. The concentration of several pro-inflammatory as well as anti-inflammatory cytokines increased significantly. IL-6, a classic endogenous, pyrogenic cytokine, increased 10 fold with this reaction demonstrating a clear physiologic response correlating with the clinical findings. Therefore, an unusual febrile reaction mediated by pyrogenic cytokines can occur in patients with AL amyloidosis exposed to a second cycle of high dose melphalan. While this reaction has not been observed in patients with multiple myeloma treated in similar fashion for unclear reasons, clinicians should be aware of this phenomenon we have termed a “melphalan recall” reaction in patients with AL amyloidosis treated with tandem cycles of HDM/SCT.
Outcome after Allogeneic Stem Cell Transplantation for Adult Acute Myeloid Leukaemia Patients Exhibiting Isolated or Associated Trisomy 8 Chromosomal Abnormality: A Survey on Behalf of the Acute Leukemia Working Party (ALWP) of the EBMT.
Authors: Chevallier Patrice , Labopin Myriam , Nagler Arnon , Ljungman Per T. , Verdonck Leo , Volin Liisa , Zander Axel , Finke Jürgen , Socie Gerard , Cordonnier Catherine , Harousseau Jean-Luc , Mohty Mohamad , Rocha Vanderson
Published: April 2020

Trisomy 8 (+8) is one of the most common chromosomal abnormality encountered in acute myeloid leukemia (AML), occurring in about 10 to 15% of cases. Currently, data assessing specifically the role of allogeneic hematopoietic stem cell transplantation (allogeneic-HSCT) in the setting of AML with +8 are still relatively sparse. The aim of this multicenter retrospective analysis was to perform a survey on overall outcomes after allogeneic-HSCT of AML patients harboring +8 as a sole chromosomal abnormality or associated with other abnormalities. We have identified 182 de novo AML patients who underwent allogeneic-HSCT between 1990 and 2007 exhibiting isolated +8 (n=136) or +8 associated to other favorable (n=8), intermediate (n=30), high-risk (n=7) or unknown (n=1) group cytogenetics reported to the EBMT. Median age was 37 years. At transplant 108 (59%) patients were in first complete remission (CR). The donor was HLA identical sibling in 115 (63%) and peripheral blood HSCT was used in 54% (n=98). Conditioning regimen was myeloablative in 82%. With a median follow-up of 48 months, 5-year non-relapse mortality (NRM), relapse rate (RR), LFS and OS were 25%, 30%, 45% and 47%, respectively. Five-years OS and LFS was not significantly different between AML patients with isolated or associated +8 (44% vs. 56% P=0.14 and 41% vs. 55%, P=0.11). In a multivariate analysis, LFS rate was improved when patients were female and transplanted in CR with an HLA identical sibling donor. LFS rate were 62% and 64% when using an HLA identical sibling donor in patients in CR1 and CR2/CR3, respectively. Isolated or associated +8 were not a risk factor for any outcomes (OS, LFS, RR, NRM). We conclude that allo-HSCT, from an HLA identical sibling donor, appears to be the treatment of choice for AML patients in CR at transplant with isolated or associated +8.
Allogeneic Transplantation (HCT) for Myelodysplastic Syndrome:Recent MDACC Experience.
Authors: Popat Uday , de Lima Marcos J , Ativitavas Touch , Rondon Gabriela , de Padua Silva Leandro , Koca Ebru , Zhang Weiqing , Alousi Amin , Anderlini Paolo , Giralt Sergio A. , Hosing Chitra M. , Jones Roy , Kebriaei Partow , Khouri Issa , Nieto Yago , Qazilbash Muzaffar H. , Andersson Borje S. , Champlin Richard
Published: April 2020

BACKGROUND: Many recent advances have occurred in the field of MDS including hypomethylating agents, lenalidomide, and WHO classification. Likewise the field of HCT has also undergone major new developments including non ablative conditioning, better supportive care, better HLA typing, selection of unrelated donors and development of reduced toxicity ablative regimens like Busulfan and Fludarabine. The role of HCT therefore needs to be redefined in light of these developments. The purpose of this study is to report our recent results.

PATIENTS AND METHODS: 89 consecutive patients with MDS as defined by WHO criteria treated at our institution between Jan 2002 and April 2008 are included in this report. There were 60 males and 29 female with a median age of 54 (23–67). There were 5(6%) patients with RA, 1(1%) RARS, 9(10%) RCMD, 22(25%) RAEB 1, 17(19%) RAEB 2, and 35(39%) therapy related MDS(t MDS). Their IPSS scores were 25(28%) patients with Intermediate 1, 49(55%) Intermediate 2, 15(17%) high. Their WPSS categories were 5(6%) patients Low, 9(10%) Intermediate, 49(55%) high, and 26(29%) very high. 51(57%) patients had a matched related donor and 38(43%) had an unrelated donor. Conditioning regimen were Flu/Bu in 56(63%) patients, Bu/Cy 1(1%), Flu/Mel 32(36%). According to HCT-CI index, the comorbidity scores were 0 in 16(18%) patients, 1 or 2 in 19(21%) and greater then 2 in 54(61%). Median time from diagnosis to transplant was 8 months (range 1–51 months).

RESULTS: With a median follow up of 28 (3–73) months, 2 year overall(OS) and disease free survival were 54%(95% CI; 42%–66%) and 52%(95% CI; 40%–64%) respectively. Cumulative incidence of non relapse mortality at 2 years was 23% (95% CI; 16%–35%). Cumulative incidence of relapse mortality at 2 years was 23% (95% CI; 15%–34%). As per WHO grouping, OS was 63%, 60%, 46% and 48% in patients with Low blast count (RA, RARS, RAMD), RAEB1, RAEB2 and t MDS( p=0.46) respectively. WPSS score was significantly(P=0.01) predictive of overall survival (see fig): 27% surviving in very high risk group, 61% in high risk group and 78% in Low and intermediate risk group. Likewise cytogenetic risk group and IPSS were significantly predictive of survival. Donor type or graft source did not predict outcome. Five patients developed primary(3) or secondary(2) graft failure. Median time to neutrophil engraftment was 13 days (8–26 days) and to platelet engraftment was 16 days (9–89 days).

CONCLUSION: These results in patients with comorbidities and with a median age of 54 years are promising. Cytogenetics and prognostic scores based on cytogenetics predict outcome after HCT.

Figure Figure
Rituximab Containing Autologous Stem Cell Transplantation May Be Curative in Mantle Cell Lymphoma for Patients in First Remission, but Not for Patients with Recurrent Disease.
Authors: Tam Constantine S , Bassett Roland , Ledesma Celina , Korbling Martin J. , Okoroji Grace-Julia , Harrell Robyn , Hsu Yvonne , De Padua Silva Leandro , Alousi Amin M. , Hosing Chitra M , Kebriael Partow , Rondon Gabriela , Giralt Sergio , De Lima Marcos , Anderlini Paolo , Popat Uday , Qazilbash Muzaffar H. , Samuels Barry , Champlin Richard E , Khouri Issa F
Published: April 2020

In order to determine if the addition of rituximab (R) had changed the outcome of autologous stem cell transplantation (AUTO) in patients with mantle cell lymphoma (MCL), we examined the outcome of 86 patients transplanted at the MD Anderson Cancer Center, in first partial or complete remission (frontline AUTO, n=50) or for relapsed / refractory disease (salvage AUTO, n=36).

<<>> Among frontline patients, 29 were transplanted in the pre-R era (AUTO-R), and 21 in the post-R era (AUTO+R). AUTO-R patients received homogenous treatment with hyper-CVAD induction and Cy- TBI conditioning. AUTO+R patients were treated with R-hyper-CVAD, hyper-CVAD and R-CHOP induction in 62%, 10% and 29% respectively, and R-Cy-TBI and R-BEAM conditioning in 43% and 52% respectively. Median age (range) at transplantation was 57 (42–66) and 56 (38–73) for the AUTO-R and AUTO+R groups respectively (p=0.91); other baseline characteristics including sex, time from diagnosis, performance status, stage, B-symptoms, B2m, LDH and MIPI were similar. Prior to transplantation, CR or unconfirmed CR (CRu) was present in 38% and 57% of AUTO-R and AUTO+R patients respectively (p=0.18), and 33 of 34 patients were negative on Gallium/PET scan. Following transplantation, CR/CRu was achieved in 93% and 100% patients respectively (p=0.50). Progression free survival (PFS) was similar between AUTO-R and AUTO+R patients for the first 2 years. However, after 2 years a clear difference emerged on the PFS curve with no relapses occurring among 9 AUTO+R patients followed between 30 and 96 months. In contrast, AUTO1-R patients experienced a continuous pattern of relapse (p=0.03 compared with AUTO+R patients). For patients remaining in remission at 2 years, no deaths had occurred in the AUTO+R group, whereas 15 of 22 AUTO-R patients had died.

<<>> The 36 salvage patients had similar baseline characteristics as frontline patients, except for the proportion of patients with chemosensitive disease (100% vs 86%, p=0.01) and negative Gallium/PET scans (97% vs 71%, p=0.01), favoring the frontline group. In contrast to the results of frontline patients, no difference was observed in salvage patients by the use of R (p=0.52 and 0.50 for PFS and OS respectively).

<<>> Autologous transplantation with rituximab has the potential to cure a proportion of patients with MCL, but only if performed during the first remission. Patients transplanted beyond first remission continued to experience a poor prognosis.

Figure Figure
Immune Reconstitution Following Myeloablative Allogeneic Hematopoietic Stem Cell Transplantation: The Impact of Expanding CD28negCD8+ T-Cells on Relapse.
Authors: Yakoub-Agha Ibrahim , Saule Pasquine , Salleron Julia , Cracco Pascale , Coiteux Valerie , Magro Leonardo , Dufosse Francoise , Bruno Benedicte , Jouet Jean-Pierre , Duhamel Alain , Dessaint Jean-Paul , Labalette Myriam
Published: April 2020

Allogeneic stem cell transplantation has become standard therapy for haematological malignancies through the positive immunologic graft-versus-leukaemia effect. Initial immune recovery relies on peripheral expansion of infused T-cells which switch to a memory-like phenotype. This study prospectively investigated whether changes in subset composition precedes late complications after myeloablative HLA-matched transplantation. Of 80 recipients, 51 experienced neither early infection nor acute graft-versus-host disease (GVHD), of whom 18 were still free of clinical complication throughout 395 – 1564 days of follow-up. Compared with this complication-free subgroup, patients who developed chronic GVHD as the only event recovered similar numbers of circulating T-cells with predominance of CD8+ T-cells lacking CC-chemokine receptor-7 and CD28 expression. Conversely, poor CD8+ T-cell recovery with diminished numbers of CD28neg CD8+ T-cells (~1/4th of that of relapse-free patients) preceded occurrence of relapse. In multivariate analysis, lower CD28neg CD8+ T-cell counts by day 60 were associated with greater risk of subsequent relapse (HR 0.33; 95% CI 0.14 - 0.76; P = 0.01). Enumeration of CD28neg CD8+ T-cells in patients without early clinical complication could assist in predicting risk of relapse and help build an algorithm for accelerating the immune recovery by reducing the immunosuppressive regimen and considering the introduction of prophylactic donor lymphocyte infusions.
Gene Expression Differences Between Fetal Liver-Derived and Adult Bone Marrow-Derived Murine Megakaryocyte Progenitors: Implications for DS-TMD.
Authors: Wieland Karen , Cantor Alan B.
Published: April 2020

About ten percent of infants with Down syndrome are born with a transient myeloproliferative disorder (DS-TMD) that spontaneously resolves within the first few months of life. Twenty to thirty percent of these infants subsequently develop acute megakaryoblastic leukemia (DS-AMKL), typically within a year or two following resolution of their DS-TMD. Recent work has shown that both DS-TMD and DS-AMKL cells harbor acquired mutations in the key megakaryocyte transcription factor GATA-1 that lead to the exclusive production of a short GATA-1 isoform (GATA-1s). The mechanism by which GATA-1s acts in DS-TMD/AMKL remains incompletely understood. Mice engineered to produce only GATA-1s exhibit hyperproliferation of fetal liver-derived megakaryocytes, but normal growth of post-natal bone marrow-derived megakaryocytes. This suggests that unique microenvironmental features of fetal liver compared to bone marrow differentially influence the effects of GATA-1s on megakaryopoiesis. In order to further understand the mechanisms involved in these stage-specific effects, we compared gene expression profiles of wild type megakaryocyte progenitors (MkPs) isolated directly from embryonic day 13.5 (e13.5) murine fetal liver (FL) and from adult bone marrow (BM). Cells were FACS sorted based on the immunophenotype lin-sca-1-c-kit+ CD41+ CD150+, which has recently been shown to mark megakaryocyte-selective progenitors. Colony forming assays of the sorted cells revealed 92–100% growth of megakaryocyte colonies in culture medium containing multilineage cytokines (SCF, IL3, IL11, GM-CSF, EPO and TPO), confirming strong enrichment for megakaryocyte lineage cells. RNA from sorted FL and BM MkPs was then used to perform Affymetrix 3′ cDNA expression microarray analysis. Expression of early-stage megakaryocyte factors, such as c-mpl, FOG1, GATA1, Runx-1 and Fli-1 were found in both sets of samples, confirming selection of megakaryocyte progenitor cells. Importantly, we observed a striking up regulation of interferon alpha (IFN alpha) inducible genes belonging to the p200 family in BM MkPs compared to FL MkPs. Gene set enrichment analyses (GSEA) confirmed broad up regulation of the IFN alpha pathway, and as well up regulation of the JAK-STAT pathway in BM MkPs compared to FL MkPs. These findings were validated by quantitative RT-PCR and in situ immunohistochemistry. Given that STAT1 is a direct GATA-1 target gene and a major downstream effector of IFN alpha signaling, we hypothesize that enhanced IFN alpha signaling in the bone marrow may compensate for potential deficiencies of STAT1 during fetal liver megakaryopoiesis in the setting of GATA-1s. Experiments are underway to test this hypothesis.
Platelets Having a W127X Mutation in GPIX Express Sufficient Residual Amounts of the GPIbα Chain to Support Ristocetin-Mediated Agglutination and Adhesion to VWF and Collagen Under Conditions of Flow.
Authors: Kanaji Taisuke , Takata Yuka , Moroi Masaaki , Seki Ritsuko , Oku Eijiro , Sano Masayuki , Nakazato Sachie , Izuhara Kenji , Sueoka Eizaburo , Imamura Yutaka , Okamura Takashi
Published: April 2020

A W127X mutation in platelet glycoprotein (GP)IX is the most common genetic defect in Japanese Bernard-Soulier Syndrome (BSS). Patients having this mutation are often misdiagnosed as having immune thrombocytopenia (ITP) because of residual expression of GPIbα on the platelet surface. The purpose of the present investigation was to determine whether the residual amount of GPIbα present on the surface of GPIXW127X BSS platelets might be sufficient to support adhesive functions of the GPIb complex. Flow cytometric analysis of platelets from our GPIXW127X BSS patient revealed 7–11% of the normal level of GPIbα (3,700 receptors/platelet), although the expression of GPIbα estimated by western blot analysis was only 1% of normal. Expression levels of other receptors such as GPIIb, GPVI, and α2 integrin were 3–4 times higher per platelet than that of normal controls–probably due to the increased size of the BSS platelets. To characterize the ability of GPIXW127X BSS platelets to bind VWF, we evaluated their agglutination in the presence of various concentrations of ristocetin, and found that ristocetin-induced agglutination was absent at 1.2 and 1.5 mg of ristocetin/ml, but present at 2.0 mg/ml. We then evaluated the ability of GPIXW127X BSS platelets to adhere under conditions of flow to immobilized VWF, type 1 collagen, and type III collagen. Whereas platelets derived from a Type I BSS patient having a TGTG deletion within the GPIbα gene in which GPIbα is completely absent (Thrombosis and Haemost 77:1055,1997) were unable to adhere to immobilized VWF under conditions of high shear, GPIXW127X BSS platelets bound well, demonstrating that the residual amount of GPIbα present GPIXW127X BSS platelets is sufficient to mediate adhesion to VWF. Finally, recent studies (J Thromb Haemost 5:797,2007) have shown that platelet adhesion to Type III collagen can be mediated by either interaction of GPIbα with VWF, or by interaction of the integrin α2β1 with collagen under high shear flow. In support of this notion, adhesion of GPIbαTGTG del BSS platelets to immobilized Type III collagen could be completely inhibited by addition of the anti-α2β1 blocking mAb, Gi9, while adhesion of GPIXW127X BSS platelets to Type III collagen was only marginally affected. Taken together, we conclude that

platelets having a W127X mutation in GPIX express residual amounts of functional GPIbα, and both GPIb-VWF and integrin α2β1-collagen interactions are physiologically important under conditions of high shear stress.

Residual expression of GPIbα in GPIXW127X platelets, therefore, likely accounts for the relatively mild bleeding phenotype in this variant form of BSS.
Angelica Polysaccharides Promotes Thrombopoiesis through the Phosphatidylinositol 3-Kinase/Akt Pathway.
Authors: Yang Mo , Chan Godfrey Chifung , Ye Jie Yu , Liu Chang
Published: April 2020

Angelica Polysaccharide is from the root of Radix Angelicae Sinensis (Chinese Danggui). Danggui has been used for centuries to treat blood-deficiency related diseases. The hematopoietic effect of Danggui may be related to its constituent, polysaccharide. However, the effects of angelica polysaccharide on hematopoietic stem/progenitor cells and megakaryocyte/platelet lineage have not been well studied. In this study, we specifically investigate the thrombopoietic effect of Angelica polysaccharide (APS) in a mouse model and its molecular mechanism. A myelosuppression mouse model (4-Gy-irradiated) was treated with APS (10 mg/kg/day) and thrombopoietin (TPO) (1 mg/kg/day). Peripheral blood cells from APS, TPO and vehicle-treated samples were counted on days 0, 7, 14 and 21. Then CFU assays were used to determine the effects of APS on the megakaryocytic progenitor cells and other lineages. Analyses of Annexin V, Caspase-3, and Mitochondrial Membrane Potential were conducted in megakaryocytic cell line M-07e. The effects of APS on cells treated with Ly294002, a Phosphatidylinositol 3-Kinse inhibitor and the effect of APS on the phosphorylation of AKT were also studied. APS as well as TPO significantly enhanced the recovery of platelet and WBC count, and bone marrow CFU-MK and CFU-GM formation (n=6). Morphological examination of bone marrows showed that APS treatment significantly increased the recovery of the megakaryocytic series. APS also increased the recovery of the body weight and organ weight (liver and spleen). However, APS didn’t have a up-regulating effect on TPO production by a ELISA assay. We further analyzed the in vitro effect of APS on CFU-MK and CFU-GM formation. APS (50 ug/ml) enhanced TPO (50 ng/ml) -induced CFU-MK (p=0.06, n=4), and CFU-GM formation (p=0.032, n=6). However, APS alone (00 ug/ml) did not show significant effect on CFU-MK and CFU-GM proliferation (n=6). The effect of APS (1–500 ug/ml) on the growth of bone marrow stromal cells was further investigated using CFU-F (fibroblastoid) assay. APS (50 ug/ml) alone showed a promoting effect on CFU-F growth (p=0.049, n=3). APS (50 ug/ml) also significantly enhanced PDGF, bFGF and VEGF-induced CFU-F formation (n=4). Moreover, the anti-apoptotic effect of APS in M-07e cells was also demonstrated by using Annexin-V, Caspase-3, and JC-1 assays. Addition of Ly294002 alone increased the percentage of cells undergoing apoptosis. However, additional of APS to Ly294002-treated cells reversed the percentage of cells undergoing apoptosis. Furthermore, addition of APS significantly increased the phosphorylation of AKT. APS promotes thrombopoiesis in a mouse model. This effect is likely to be mediated by the PI3K/AKT pathway.
Blocking of IL-21 Signal Attenuates Graft-Versus-Host Disease but Not Graft-Versus-Leukemia Effect in a Mouse Model
Authors: Meguro Akiko , Ozaki Katsutoshi , Oh Iekuni , Matsu Haruko , Hatanaka Keiko , Tatara Raine , Leonard Warren J , Sato Kazuya , Ozawa Keiya
Published: April 2020

Immunological effects of interleukin-21 (IL-21) on T-, B-, and NK-cells have been reported, but the role of IL-21 in graft-versus-host disease (GVHD) and graft-versus-leukemia (GVL) remains obscure. Here, we demonstrate that splenocytes from IL-21R−/− mice show attenuated GVHD symptoms as compared to splenocytes from wild type mice. In addition, the survival of mice receiving IL-21R−/− splenocytes is significantly longer than of those receiving wild type splenocytes. Both types of splenocytes appear to efficiently eliminate leukemic cells, suggesting the retention of GVL effects. Our results suggest that CD4 T-cells augment GVHD by autocrine stimulation via IL-21, and that production of IL-17 is downregulated in IL-21R−/− splenocyte recipients. To develop a treatment model in mice, we generated a decoy receptor for IL-21, in which most of the intracellular domain is deleted. Mice receiving bone marrow cells transduced with this decoy receptor survived longer than those receiving cells transduced with an empty retrovirus.These results suggest that IL-21 promotes GVHD and that the blockade of IL- 21 signaling might be a useful treatment for GVHD, without causing loss of the GVL effect.
Immune Response of Human Polymorphnuclear Neutrophils to Aspergillus Fumigatus
Authors: Loeffler Juergen , Wozniok Iwona , Mezger Markus , Einsele Hermann
Published: April 2020

Invasive fungal infections with Aspergillus fumigatus show an increasing incidence due to the growing number of severely immunocompromised patients. Polymorphonuclear neutrophils (PMNs), as part of the innate immune system, are key players in antifungal immune responses, recognizing fungal pathogens at an early step of infection. Besides phagocytic mechanisms, PMNs kill pathogens by the release of reactive oxygen species (ROS). Human PMNs were isolated from blood of healthy donors using Biocoll separation. Cells were co-cultivated with conidia, germlings and hyphae of the clinical strain ATCC 9197. The oxidative burst was determined in a kinetic measurement quantifying dichlorfluorescein production. Chemotaxis was analyzed by transwell assays. Furthermore, total RNA was extracted and gene expression profiling was performed using Affymetrix U133Plus2.0 arrays. We could demonstrate that A. fumigatus represents a strong stimulus releasing ROS, depending on the morphotype; germlings revealed high ROS release, whereas resting conidia showed low stimulation capacity. PMNs actively tracked germlings and directly attached to fungi as demonstrated by real-time microscopy. Transwell assays revealed that chemotaxis of PMN is strongly dependent on the fungal morphotype; PMN showed only weak chemotaxis in the presence of conidia, whereas in the presence of germlings, high chemotactic activity was achieved. Furthermore, after 6h co-cultivation of PMNs and A. fumigatus germ tubes, 195 (1.273) genes showed an at least 4fold (2fold) altered gene expression. We observed upregulation of genes (hemoxygenase, heat shock 70kDa protein, HSPA8, HSPA1B, HSP90AB1, Ferritin) involved in self-protection against radicals. Additionally, genes involved in inflammatory responses (IL-8, CCL3, CXCL2, IL1RN) were significantly upregulated. Luminex analysis was performed for TNF-a, IL-12, GM-CSF, IFN-γ, IL-6, IL-8, IL-10 and IL-1b to identify secreted cytokines, thereby confirming array data. In conclusion, A. fumigatus had substantial effects on the activation of human PMNs. Various defence strategies were initiated, including phagocytosis, ROS release and mobilization of other immune effector cells by secretion of chemoattractants.
Familial Hemophagocytic Lymphohistiocytosis Type IV: Possible Founder Effect in Syntaxin 11 Gene Common Mutation Endemic to Turkish Population
Authors: Balta Gunay , Caliskan Umran , Guler Elif , Patiroglu Turkan , Oner Ahmet F. , Gurgey Aytemiz
Published: April 2020

Familial Hemophagocytic Lymphohistiocytosis (HLH) is a rare, autosomal recessive, if untreated, fatal disorder of early childhood characterized mainly by persistent fever, hepatocyplenomegaly, cytopenia, hypertriglyceridemia, hyperferritinemia, defective natural killer cell activity and hemophagocytosis in bone marrow, lymph nodes, liver and spleen. Familial HLH is a genetically heterogeneous disease; at least four sub-types (I–IV) associated with three genes and one chromosomal locus (9q21.3-q22) have been reported in the disease to date. A number of mutations in both Perforin and Munc13– 4 genes have been described in different populations whereas only 3 different types of mutations in Syntaxin 11 gene have, thus far, been described in a few families solely from Turkish population. Therefore, elucidation of the mutation spectrum of the Syntaxin 11 gene mostly depends on the identification of its genetic defects in this population. In this communication, we present our findings on Syntaxin 11 gene which is responsible for the familial HLH type IV. A total of 84 potentially primary HLH patients selected on the basis of consanguinity and/or family history and/or relapse were included in the study. Patients and family members were studied by linkage analysis for all 4 described gene loci and homozygosity for Syntaxin 11 gene was observed in 20 patients from consanguineous families. Sequencing of these patients for the complete coding and flanking sequences of Syntaxin 11 gene revealed molecular defects in only 5 patients (2M/3F) with the ages ranging from 2 months to 3 years. Contrary to the heterogeneity in the mutation spectrum of hereditary disorders generally observed in Turkish population, a single type of homozygous mutation (c369–370delAG, c374–376delCGC) was identified in these 5 unrelated patients who had a family history of at least one deceased sibling except one. Screening of the family members revealed that the obligate carriers and some of the siblings were heterozygous for the mutation. This 2 plus 3 nucleotide deletion in exon 2 leads to frameshift and premature termination of translation 59 altered aminoacids downstream (Val124fsX60). Haplotype analysis indicated that these patients who were coming from different parts of Turkey may have a common ancestor (founder effect). Besides confirmation of the “Turkish population endemic” profile of the Syntaxin 11 gene mutations, the findings provided here elucidate the single/common mutation characteristic of the gene which is extremely important in the molecular diagnosis of the disease. This study was supported by TUBITAK (Project No: 105S386; SBAG-3193).
Functional Characterization of Signaling Events Underlying Immunostimulatory and Immunosuppressive Activity of Macrophages.
Authors: Kloss Mercedes , Kanz Lothar , Salih Helmut R , Krusch Matthias
Published: April 2020

Macrophages (Mo) play an important role in combating infectious pathogens and regulating adaptive immune responses. Their reactivity is, among others, guided by pattern recognition receptors including Toll like receptor 4 (TLR4), which recognizes bacterial lipopolysaccharide (LPS). After stimulation, TLR4 can associate with different adaptor proteins like e.g. MyD88 or TRIF leading to the activation of signaling molecules like MAPK or NF-κB. However, stimulation with LPS may result in equally impressive immunostimulatory and immunosuppressive Mo responses. The signaling events responsible for this diametrical behavior of different Mo subsets are yet not fully understood. To study the effects of LPS on Mo we differentiated monocytes of healthy donors with GM-CSF and M-CSF into pro-inflammatory type 1 (Mo1) or anti-inflammatory type 2 (Mo2) subsets, respectively. As expected, upon LPS stimulation the pro-inflammatory Mo1 secreted low levels of the immunosuppressive cytokine IL-10 and high levels of IL-12, while the anti-inflammatory Mo2 produced neglectable amounts of IL-12 but high levels of IL-10. To elucidate the underlying molecular mechanisms we performed gene array analyses with Mo1 compared to Mo2 from three independent donors. Interestingly, among over 50 genes involved in TLR4 signaling, only TLR4 itself and its adaptor protein MyD88 were found to be differentially regulated: Within the immunosuppressive Mo2 subset, MyD88 was significantly downregulated, while a significant upregulation of TLR4 was observed compared to the immunostimulatory Mo1 subset. The differential expression pattern were further confirmed on protein level by western blot. Since TLR4 stimulation leads to MyD88-dependent activation of the MAPK JNK, we investigated whether JNK was differentially involved in cytokine production of Mo1 and Mo2. Addition of a specific JNK inhibitor (SB600125) concentration dependently reduced TLR4-induced IL-10 but enhanced IL-12 production in Mo1, pointing to an immunoinhibitory role of JNK in Mo1. In contrast, inhibition of JNK did not alter the secreted levels of IL-12 or IL-10 in Mo2, indicating that rather MyD88-independent pathways are responsible for the anti-inflammatory behavior of Mo2. In conclusion, our data suggest that the opposite regulation of the adaptor molecule MyD88 in pro- and anti-inflammatory Mo subsets is responsible for the fact that a single, non-polymorphic receptor like TLR4 can mediate both pro- and anti-inflammatory immune responses in Mo.
Patient-Reported Fatigue Scores in Cancer Patients with Chemotherapy-Induced Anemia Treated with Erythropoietic-Stimulating Agents (ESAs).
Authors: Burton Tanya , Hoaglin David , Larholt Kay , Apgar Elizabeth , Pashos Chris , Bookhart Brahim , Corral Mitra , Piech Catherine Tak , McKenzie R. scott
Published: April 2020

Background: Clinical trial evidence has demonstrated greater anemia-related fatigue at lower hemoglobin (Hb) levels in ESA-treated patients with chemotherapy-induced anemia (CIA). To understand these patient-reported outcomes (PROs), during ESA treatment in a real-world setting, data were analyzed from the Dosing and Outcomes Study of Erythropoiesis-Stimulating Therapies (D.O.S.E.) Registry, an ongoing, prospective, observational study.

Methods: Patients included for analysis were required to have a baseline Hb prior to ESA administration and at least two ESA administrations. PROs were measured using the Functional Assessment of Cancer Therapy: Fatigue subscale (FACT-F), a 13-question measure of anemia-associated fatigue with lower scores indicating greater fatigue in patients with CIA. Patients were to have completed up to three FACT-F questionnaires within seven days of a post-baseline Hb level. If multiple Hb values were identified, the Hb value closest to the FACT-F assessment was reported. The association between FACT-F scores and Hb levels was tested in a repeated-measures analysis. Mean FACT-F scores were calculated for specified Hb ranges.

Results: 740 CIA patients from 63 sites were identified between 12/03 and 7/08. 1088 concurrent FACT-F and Hb level assessments were available during ESA treatment. Mean FACT-F scores were lower when patients had lower Hb levels (table). The association between FACT-F scores and Hb levels showed a significant positive trend (p<.0001).

Hb ≤ 8.0 g/dL 8.01–9.0 g/dL 9.01–10.0 g/dL 10.01–11.0 g/dL 11.01–12.0 g/dL n (concurrent FACT-F and Hb assessments) 19 63 212 336 458 Mean FACT-F score (SD) 21.26 (11.01) 22.95 (12.44) 25.64 (12.40) 26.84 (11.70) 29.34 (12.04)

Conclusions: This analysis of FACT-F assessments in CIA patients suggests that lower Hb levels were associated with greater anemia-related fatigue. These findings are consistent with clinical trial evidence and provide observational PRO data to patients and providers regarding anemia-related fatigue during ESA treatment.
Unrelated Cord Blood Transplantation (UCBT) for Transfusion- Dependent Thalassemia a CIBMTR Audited Retrospective Analysis of 30 Consecutive Patients from a Single Center
Authors: Jaing Tang-Her , Wang Brian , Gjertson David , Law Ping , Petz Lawrence , Chow Robert
Published: April 2020

UCBT offers a cure for thalassemia and has advantages as a stem cell source because the requirement for an HLA match between donor and recipient is less stringent. Previous reports of UCBT for thalassemia have yielded transplant related mortality (TRM) that was quite high and it is known that pre-freeze total nucleated cell (TNC) dose is critical for UCBT. With strategies that maximize TNC dose – using non-red cell reduced but plasma depleted (PD) CB, no post-thaw wash (NW), and double cord transplantation (DCT) when necessary – we have achieved promising results with UCBT in young patients with transfusion-dependent β-thalassemia. Between 10/2003 and 5/2008, 39 CB products were infused after Bu/Cy/ATG myeloablation into 30 pediatric thalassemia major patients (9 DCT) using NW PD CB exclusively. Patient status: 21 Pesaro class−1, 8 Pesaro class−2, and 2 unknown. Median age was 5 years (range 1–14 years) with a median weight of 18 kg (range 11–37 kg). The data was audited by the transplant center (TC) and on-site by CIBMTR (98.5% accuracy for Chang Gung). Median pre-freeze TNC cell dose was 10.9×107/kg, and median pre-freeze CD34+ cell dose was 4.0×105/kg. The number of CB products HLA ABDR matched were 4, 11, and 24 for 6/6, 5/6, and 4/6 or fewer matches. No significant adverse events were observed despite major ABO incompatibility in some cases and forgoing post-thaw wash. Cumulative incidences of donor-derived (determined by chimerism studies) neutrophil (ANC500), platelet 20K and 50K (plt 20K & 50K) engraftment were 96±11%, 92±11% and 92±12%, and median times to ANC500, plt 20K and 50K engraftment were 17.5 days (range 11–26), 49.5 days (range 28–135) and 63.5 days (range 35–203), respectively. Grade III/IV acute GvHD occurred in 40±9%, and extensive chronic GvHD occurred in 4±4% of the patients. Transplant related mortality were 10±5% at 100 days and 13±6% at 1 year. Overall survival at 1 and 3 years were 87±6% and 82±8%, while disease-free survival at 1 and 3 years were 85±7% and 78±9%. Five patients expired including 1 death prior to day +10 due to traumatic head injury. Mean and median follow up times were 24 and 16 months respectively (range 0.3–58 months) as of August 2008. To our knowledge, this is the largest single-institutional UCBT series for thalassemia, and shows the favorable clinical results that are attainable when TNC dose is optimized with PD CBU, no post-thaw wash and DCT when necessary.
Participation in a Clinical Trial Does Not Impact Outcome in Pediatric Acute Lymphoblastic Leukemia
Authors: Koschmann Carl , Hawkins Doug S , Thomson Blythe
Published: April 2020

Background: Acute lymphoblastic leukemia (ALL) is the most common pediatric malignancy. Over the past four decades, the outcome for pediatric ALL has rapidly improved secondary to the participation of children on clinical trials, resulting in standardization of the treatment. Some have argued that the best outcome results from participation in a pediatric clinical trial. To address the potential benefits and barriers to participation in a clinical trial, we analyzed clinical trial participation in pediatric ALL at a large single institution.

Methods: We evaluated 322 ALL patients < 22 years old at diagnosis who receive their initial therapy at Seattle Children’s Hospital Regional Medical Center (S-CHRMC) between January 1997 and December 2005. Using a retrospective chart review, we analyzed the following variables: study participation (SP) or non-participation (NP) in an ALL therapeutic study, gender, race, patient immunophenotype, risk group (standard risk (SR), high risk (HR) or infant), home state, and distance of primary residence from S-CHRMC. S-CHRMC is the largest pediatric cancer center in the Pacific Northwest, with referrals from Washington, Alaska, Montana, Idaho and Wyoming. Events were defined as relapse or death from any cause.

Results: The overall 5 year event free survival (EFS) was 78% (+/− 2.5%). 157 patients participated in a treatment research study (49%). Only risk group was associated with EFS. SP and NP had similar EFS. Gender, race, immunophenotype, home state, or distance from primary residence were not associated with outcome. There was no difference in study participation by gender, race, home state, or distance of primary residence. There were trends to increased participation in SR vs HR (54% vs 35%, p value 0.15) and B lineage vs T lineage (50% vs 35%, p value 0.11).

Variable n 5 year EFS p value Study participation No 165 76 (+/− 3.9) 0.47 Yes 157 81 (+/− 3.3) Risk group SR 175 85 (+/− 3.0) HR 132 73 (+/− 4.2) (0.038 to SR) Infant 15 59 (+/−12.9) (0.011 to SR)

Discussion: Participation in a research study for treatment of pediatric ALL was not associated with improved outcome in our large single institution series. Study participation was not different by clinical features, including distance to the primary residence. Strict standardization of treatment for all patients may contribute to the similar outcome for SP and NP.
Altered Erythroid and Megakaryocytic Differentiation in Mice Expressing a Unique Chromatin Remodeling Domain of Erythroid Krüppel-Like Factor (EKLF)
Authors: Jansen Valerie M. , Abramova Tatiana , Shim Eun-Hee , Ramachandran Shaji , Desgardin Aurelie , Parekh Vishwas , Jane Stephen , Cunningham John M
Published: April 2020

The zinc finger-encoding transacting factor EKLF binds key regulatory elements of many erythroid-specific genes, and is essential for definitive erythropoiesis. Mice lacking this factor (EKLF−/−) die of anemia by E15.5 of gestation, failing to activate β-globin gene transcription, and demonstrating a block in the erythroid differentiation program at the primitive erythroblast stage. In contrast, megakaryocytic progenitors are amplified in EKLFnull embryos, with increased Fli-1 gene expression (a marker of early megakaryocytic differentiation), consistent with the idea that EKLF modulates the megakaryocyticerythroid (M-E) differentiation switch. We have demonstrated that an amino terminal mutant of EKLF (Δ221EKLF), is required to induce chromatin remodeling at the β-globin promoter in an EKLF-null erythroid cell line, but additional amino terminal sequences are required for initiation of β-globin gene transcription (Brown et al., 2002). To evaluate the role of this chromatin remodeling (CR) domain in erythroid and megakaryocytic differentiation in vivo, we have generated a knock-in allele of EKLFCR allele. Similar to EKLF-null embryos, mice homozygous for this mutant allele die of anemia by E15.5 of gestation. In contrast to erythroid cells lacking EKLF, EKLFCR/CR progenitors demonstrate

appropriate binding of the CR encoding domain to all EKLF-regulatory sequences; a block in erythropoiesis at a more a mature stage in differentiation a chromatin architecture and histone modification pattern at erythroid-specific genes that recapitulates the events observed in EKLF+/+ erythroblasts at a similar stage of erythroid ontogeny; a failure of terminal erythroid gene transcription.

Examining the role of EKLFCR in megakaryopoiesis, we observed

inhibition of megakaryocytic progenitor amplification in EKLFCR/CR fetal hematopoietic cell populations when compared to EKLF-null embryos; loss of Fli-1 gene expression in EKLFCR expressing cells; binding of the EKLFCR mutant protein to the Fli-1 promoter with inhibition of gene transcription; a repressed chromatin architecture at megakaryocytic gene loci.

In contrast to these results, mice homozygous for a knockin allele encoding the zinc finger DNA binding domain alone (Δ253EKLF), a region shown previously to be sufficient for chromatin remodeling in vitro, demonstrate erythroid and megakaryocytic phenotypes that resemble those observed in EKLF-null hematopoietic progenitors. Taken together, our results suggest strongly that the unique EKLFCR domain is necessary and sufficient to modulate the chromatin-specific roles of EKLF at erythroid- and megakaryocytic-specific loci in definitive hematopoietic cells in vivo.
Lymphoma and Whole Body CT Scan Mortality Risk
Authors: Shenoy Pareen J. , Gude Tulasi , Sinha Rajni , Flowers Christopher R.
Published: April 2020

Background: Recent concerns regarding the risks of cancer and cancer-related death due to radiation from computed tomography (CT) scans (Brenner, NEJM 2007) and the lack of data demonstrating a survival advantage for surveillance CT scans following lymphoma therapy have raised questions regarding their benefit. We compared the radiation-related lifetime cancer incidence (LCI) and mortality risks (LCMR) associated with CT scans for staging and surveillance of non-Hodgkin lymphoma (NHL) and Hodgkin lymphoma (HL) with the cumulative probability of lymphoma death (CPLD) during monitoring.

Methods: Using 2008 NCCN Guidelines for HL and NHL, we estimated that the average number of scans a patient would need during 5 years of treatment and follow-up would be 10. We utilized published estimates of the cumulative organ-specific radiation doses from full-body CT scans (Brenner, Radiology 2004) and calculated the LCI and LCMR using sex-, age-, and organ-dependent cancer risks per 0.1Gy provided by the Biological Effects of Ionizing Radiations VII report. Site-specific cancer risks were summed to yield the overall LCI and LCMR as described by Brenner (Med Phys 2001). Surveillance, Epidemiology, and End Results (SEER) data were used to identify cases between 2000 and 2005 from 17 SEER Registries and calculate CPLD for specified cohorts.

Results: LCI and LCMR were lower for males and were markedly less than the CPLD at 5 years for most patients, but relevant for younger women with HL given their low risk of HL death (Table 1). The relationship between LCMR and the number of CT scans is shown in Figure 1. Additional implications of surveillance strategies on specific cancer risks will be presented.

Conclusion: While the LCMR from CT scans is small compared to lymphoma-related deaths for most subgroups, these data should be discussed with patients in formulating plans for surveillance following lymphoma therapy.

Table 1 LCI and LCMR due to CT scans compared with CPLD for lymphoma subtypes by age at diagnosis and gender

Lymphoma Subtype (Age at Diagnosis) LCI from CT* LCMR from CT* 5-year Cumulative Probability of Lymphoma Death** Males Females Males Females Males Females * per person exposed to the cumulative radiation dose associated with 10 full-body CT scans during 5 years from diagnosis ** Age group studied were HL (20–29 & 80–84), DLBCL (60–79), FL (60–79), NHL (60–79) NHL (70 yrs) 0.0044 0.0057 0.0032 0.0044 0.31 0.28 FL (70 yrs) 0.22 0.18 DLBCL (70 yrs) 0.39 0.37 HL (20 yrs) 0.0133 0.0242 0.0070 0.0108 0.07 0.07 HL (80 yrs) 0.0022 0.0030 0.0019 0.0026 0.47 0.55

Figure 1 LCMR due to CT scans by age at diagnosis of lymphoma and gender Figure 1. LCMR due to CT scans by age at diagnosis of lymphoma and gender