Liver GVHD Research Articles

Comparison of tacrolimus vs. cyclosporine in pediatric hematopoietic stem cell transplantation for thalassemia.

Graft-versus-host disease (GvHD) is one of the leading causes of morbidity and mortality in patients undergoing allogeneic HSCT, and effective prevention of GvHD is critical for the success of the HSCT procedure. Calcineurin inhibitors (CNI) have been used for decades as the backbone of GvHD prophylaxis. In this study, the efficacy and safety of Cyclosporine A (CsA) and tacrolimus (TCR) were compared in pediatric HSCT for thalassemia. This is a retrospective analysis of 129 pediatric patients who underwent HSCT with the diagnosis of thalassemia at Medicalpark Göztepe and Antalya Hospitals between January 2017 and December 2020. Despite the GvHD prophylaxis, grade II-IV acute GvHD developed in 29 patients. Of these patients, 12 had only gut, 10 had only skin, 6 had combined gut and skin, and one had only liver GvHD. Fifteen of these 29 patients were in the CsA group, and 14 of them were in the TCR group. There was no significant difference between the groups in terms of acute GvHD occurrence, GvHD stage, or involvement sites. In terms of CNI-related toxicity, neurotoxicity in 15 (CsA n = 9, TCR n = 6) and nephrotoxicity in 18 (CsA n = 4, TCR n = 14) patients were observed. While there was no difference between the two groups in terms of neurotoxicity, more nephrotoxicity developed in patients using TCR (p = .013). There was no significant difference between the groups in terms of engraftment syndrome, veno-occlusive disease, CMV reactivation, PRES, or graft rejection. Regarding GvHD, there was no difference in efficacy between TCR and CsA usage. Patients taking TCR experienced noticeably higher nephrotoxicity in terms of adverse effects. This difference should be considered according to the patient's clinical situation while choosing a CNI.

Recipient Resident Macrophage Plays a Crucial Role in Acute Graft-Versus-Host-Disease after Allogeneic Stem Cell Transplantation By Regulation of Damage-Associated Molecular Pattern Phagocytosis

Introduction Evidence shows allogeneic hematopoietic stem cell transplantation (alloHSCT) provides life-sustaining therapy for many hematologic malignancies. Unfortunately, in many cases it is complicated by acute-graft-versus-host disease (aGVHD), a life-threatening sequela. Most interstitial tissues contain populations of resident macrophages. Multiple studies have investigated the role of infiltrating macrophages in aGVHD, but little is known about the role of resident macrophage in aGVHD. To address this issue, we adopt a LysM cre/cre-Hif1a flox/flox mice (KO)as host mice with abnormal macrophage whose host resident macrophage remains unchanged early after allo-HSCT. KO mice received alloHSCT after Total body irradiation (TBI) from wild-type(WT) mice of BALB/c background. The neutrophil and circulating monocyte/macrophage were replaced by WT cells and the resident macrophage remains unchanged after this procedure. The grade of aGVHD and survival was observed. Methods Recipient mice are on the background of C57BL/6(H-2 b), and donor mice are wild-type BALB/c (H-2 d ) mice. All mice used were 8-12 week old. All animal experiment protocols were approved. LysM cre/cre (CRE) mice were taken as control recipients. Mice were exposed to TBI split into two doses separated by 4h. KO and CRE C57/BL mice received 9 Gy TBI. 18h after irradiation, mice of two groups were injected i.v with 1×10 7BM cells and 1.5×10 7 splenocytes isolated from WT BALB/c mice. aGVHD score and survival are monitored daily. Isolation of Kupffer subsets was performed as CD45 +CD11b + Ly6G -F4/80 +, which from KO and CRE mice of GVHD model were taken to do single-cell sequencing assay. Survival curves were plotted by the Kaplan-Meier method. The statistical significance between group means was determined using t-test by Graphpad prism9.5. The level of significance was set at P<0.05. Results The mean survival of KO mice is 18.6 days (95%CI 14.06-22.6) post alloHSCT, while the mean survival of control mice is 37.6 days (95%CI 29.7-45.6)(n=8)( P <0.001). Weight loss (% of baseline weight on the day of TBI) starts to differ on day 14 post-HSCT, with the KO group of 101.7 and Control group of 84.9 ( P <0.05). GVHD score starts to differ on day 10 post-HSCT, with the KO group of mean score 5.1 and control group 3.9 ( P <0.05)(n=8) (Figure1). The GVHD score remains statistically different till day 24 post-HSCT. On day 24, all KO mice die while one control mouse live up to 58 days. The liver function presented by ALT is different on day 14 post-HSCT, KO group with mean ALT 209.0 U/L is higher than Control group whose mean ALT is 64.9U/L(n=4). On day 21, Liver GVHD pathology score of the KO group is 7, compared to 5.2 of the control group(n=6)( P<0.05). Thus, KO mice present with a more severe aGVHD phenotype than CRE mice. Macrophage is known to have deficient phagocytosis ability when lacking Hif1-a, which is also confirmed through phagocytosis assay in our lab by FITC marked particles based on melamine resin ( P <0.05). We hypothesize the disability of phagocytosis of recipient resident macrophage renders more DNA debris (also known as DAMP) generated from conditioning, leading to susceptibility of aGVHD, we applied Dnase I to mice from day4 to day14 post-HSCT. The survival of KO mice after injection of Dnase I, with a mean survival of 39.9 days, is better than KO mice with vehicle injection ( P <0.05). After Dnase I injection, KO group, compared to 63.5 days mean survival of the control group, there is no statistical difference ( P=0.113). In order to look into more mechanisms, CD45 + cells from the liver of KO and CRE mice with GVHD model 21 days post-HSCT were isolated to do single-cell sequencing. KO mice revealed more T cell ratio than CRE mice. GO analysis revealed several abnormalities including regulation of lymphocyte activation. Conclusions We build GVHD model on LysM cre/cre-Hif1a flox/flox and LysM cre/cre mice, both adopt alloHSCT from wild-type BALB/c mice. After HSCT, both groups are replaced with wild-type neutrophils and circulating monocyte/macrophages, leaving resident macrophages different. KO mice present with exacerbated GVHD phenotype than CRE mice after alloHSCT. This difference could be mitigated by Dnase I injection. Thus, the resident macrophage of the recipient plays a crucial role in aGVHD occurrence, which is partly by regulation of DAMP phagocytosis.

Iron Dys-Homeostasis Contributed to Liver Gvhd after Allogeneic Hematopoietic Stem Cell Transplantation

Graft-versus-host disease (GVHD), which results from immunological attack to recipient organs or tissues (such as the skin, liver and gut) by donor allogeneic T cells, limits the success of allogeneic hematopoietic stem cell transplantation (allo-HSCT). A correlation of high serum ferritin levels with increased mortality after allo-HSCT has been suggested by several analyses. While the role of iron dys-homeostasis in the risk of GVHD remains unclear. Whether iron dys-homeostasis contributes to the development of GVHD or iron dys-homeostasis is only a result of severe GVHD remains unknown. To assess the status of iron metabolism during GVHD following allo-HSCT, we established aGVHD mouse model by transplantation of T cell-depleted bone marrow (TCD BM) with or without splenic T cells from C57BL/6 (B6) mice to BALB/c mice after being lethally irradiated. By 28 days post-transplantation, when aGVHD stable developed, the serum iron level was significantly higher in aGVHD group compared with control. Meanwhile, ferrous-ion load in aGVHD-involved liver was also increased, while the total iron load did not differ significantly in aGVHD-involved liver compared with control. The ferrous-ion load as well as the total iron load in intestine, lung and spleen were comparable between aGVHD group and control group. To further determine the dysregulation of iron metabolism during liver aGVHD, we detected the expression levels of pivotal iron regulatory genes in liver and duodenum. The expression levels of transferrin receptor 1 ( Tfr1) and divalent metal transporter 1 ( Dmt1) were significantly upregulated in aGVHD-involved liver, which contributed to ferrous-ion overload in the liver aGVHD. Besides, Ferroportin 1 was down-regulated in aGVHD-involved duodenum, which is the result of the upregulated hepcidin ( Hamp1) in liver. Our data suggested that the increased serum iron in aGVHD was not related to iron absorption by duodenum, but may be associated with ineffectively use of iron, which was caused by the impaired marrow function after myeloablative conditioning. To explore whether ferrous-ion overload in liver would lead to induction of reactive oxygen species (ROS) via the Fenton reaction, and eventually results in liver damage during aGVHD, we detected the activity of superoxide dismutase (SOD) and the load of thiol and lipid peroxidation product, malondialdehyde (MDA). The activity of SOD and thiol load both decreased significantly in aGVHD-involved liver, while the serum MDA level increased in aGVHD group, indicating that iron dys-homeostasis may aggravate liver GVHD damage by induction of lipid peroxidation injury. Finally, to explore how iron overload affect GVHD, we fed recipient mice with high-iron diet and low-iron diet since three weeks before transplantation, respectively. The survival rates and GVHD score were significantly worse in high-iron diet group compared with normal-iron diet group. Interestingly, low-iron diet didn't show survival benefit compared with the normal-iron diet. This suggests that excessive iron deficiency is not beneficial for preventing and treating GVHD. Thus, we applied apo-transferrin (apo-Tf) administration to receipt mice, which can render the poorly soluble non-transferrin bonding iron (NTBI) into a soluble transferrin-bound form, and prevent iron from being rapidly uptake by the liver. The administration of apo-Tf alleviated the organ damage during aGVHD, and reduced the infiltration of CD4 +IFN-γ +T cells, CD4 +IL17 +T cells and CD8 +IL17 +T cells in target organs such as intestine, liver and lung. These data suggested that ferrous-ion overload leads to the increase of NTBI, which aggravate GVHD tissue damage. Our findings reveal a novel mechanism of iron dys-homeostasis during the development of liver aGVHD, which may reveal promising therapeutic targeting in the prevention and treatment of liver aGVHD.

Humanized anti-CD25 monoclonal antibody as a salvage therapy for steroid-refractory acute graft-versus-host disease after hematopoietic stem cell transplantation

Objective: To investigate the efficacy of humanized anti-CD25 monoclonal antibody for steroid-refractory acute graft-versus-host disease (SR-aGVHD) in allogeneic hematopoietic stem cell transplantation (allo-HSCT) recipients. Methods: A total of 64 patients with SR-aGVHD between June 2019 and October 2020 in Suchow Hopes Hematology Hospital were enrolled in this study. Humanized anti-CD25 monoclonal antibodies 1 mg·kg(-1)·d(-1) were administered on days 1, 3, and 8, and then once per week according to the disease progression. Efficacy was assessed at days 7, 14, and 28 after humanized anti-CD 25 treatment. Results: Of the 64 patients with a median age of 31 (15-63) years, 38 (59.4%) were male and 26 (40.6%) were female. The overall response (OR) rate of the humanized CD25 monoclonal antibody in 64 patients with SR-aGVHD on days 7, 14, and 28 were 48.4% (31/64), 53.1% (34/64), and 79.7% (51/64), respectively. Liver involvement is an independent risk factor for poor efficacy of humanized CD25 monoclonal antibody for SR-aGVHD at day 28 (OR=9.588, 95% CI 0.004-0.291, P=0.002). The median follow-up time for all patients was 17.1 (0.2-50.8) months from the start of humanized CD25 monoclonal antibody therapy. The 1- and 2-year OS rates were 63.2% (95% CI 57.1% -69.3%) and 52.6% (95% CI 46.1% -59.1%), respectively. The 1- and 2-year DFS rates were 58.4% (95% CI 52.1% -64.7%) and 49.8% (95% CI 43.4% -56.2%), respectively. The 1- and 2-year NRM rates were 28.8% (95% CI 23.1% -34.5%) and 32.9% (95% CI 26.8% -39.0%), respectively. The results of the multifactorial analysis showed that liver involvement (OR=0.308, 95% CI 0.108-0.876, P=0.027) and GVHD grade Ⅲ/Ⅳ (OR=9.438, 95% CI 1.211-73.577, P=0.032) were independent risk factors for OS. Conclusion: Humanized CD25 monoclonal antibody has good efficacy and safety for SR-aGVHD. This study shows that SR-aGVHD with pretreatment grade Ⅲ/Ⅳ GVHD and GVHD involving the liver has poor efficacy and prognosis and requires early intervention.

Advantage of First-Line Therapeutic Drug Monitoring-Driven Use of Infliximab for Treating Acute Intestinal and Liver GVHD in Children: A Prospective, Single-Center Study.

The high serum concentrations of TNF-α characterize acute graft-versus-host disease (aGVHD), for which infliximab treatment may be beneficial. In 28 pediatric patients, four doses of 10 mg/kg infliximab every seven days were administered after steroid failure (Standard Group, n = 14) or as a first-line therapy (Early Group, n = 14). Population pharmacokinetic analyses and evaluation of serum cytokines were performed. After two months of treatment, complete response in gastrointestinal and liver aGVHD was achieved in 43% and 100% of patients in the Standard and Early groups, respectively. During follow-up, four patients in the Standard Group (but none in the Early Group) experienced an aGVHD recurrence. Viral infections occurred more frequently in the Standard Group after the fifth dose. Infliximab clearance did not differ between groups or according to treatment outcome for each organ involved in aGVHD, whereas serum levels of cytokines significantly differed. Therefore, present findings show that use of first-line, TDM-driven infliximab to treat aGVHD in children may result in better clinical outcomes and tolerability, with a different pattern of cytokines generated according to the moment of beginning of treatment.

Correlative factors of ocular surface lesions after allogeneic hematopoietic stem cell transplantation: A retrospective study.

Ocular graft-versus-host disease (oGVHD) is one of the complications after allogeneic hematopoietic stem cell transplantation (HSCT), which impairs the quality of life and may indicate poor prognosis. In this retrospective study, the aim was to investigate the characteristics of ocular surface after HSCT, and analyze the risk factors related to the severity of ocular surface lesions. 248 post-HSCT patients were enrolled in this retrospective study. Subjects were divided into no lesion group, mild lesion group and severe lesion group, according to the severity of ocular surface lesions. The correlations between grades of ocular surface lesions and gender, age, primary disease, donor source, human leukocyte antigen (HLA) type, kinship, donor-recipient relationship, blood type, source of stem cell and systemic GVHD were analyzed. The median scores of corneal epitheliopathy, lid margin lesions and meibomian gland loss were 3, 6 and 2 points, respectively. The grade of corneal epitheliopathy was related to donor source (P<0.001), kinship (P=0.033), HLA-matching (P<0.001), and systemic GVHD (P=0.007), especially oral GVHD (P<0.001) and liver GVHD (P=0.002). The grade of lid margin lesions was related to donor source (P=0.019), HLA-matching (P=0.006), and systemic GVHD (P=0.013), especially skin GVHD (P=0.019) and oral GVHD (P=0.019). The grade of meibomian gland loss was related to age (P=0.035) and gastrointestinal GVHD (P=0.007). The grade of corneal epitheliopathy after HSCT was related to the lid margin lesion score (P<0.001). The occurrence and development of ocular GVHD are mostly accompanied by the history of systemic GVHD. While in few cases, ocular surface lesions related to GVHD can be observed prior to the rejection of other tissues and organs. Severe corneal epitheliopathy occurs in patients with severe lid margin lesions in ocular GVHD. The lesions of corneal epithelium and lid margin are milder in HLA partially matching transplantation.

Multicenter Phase I Study of Post-Transplant Low-Dose Inotuzumab Ozogamicin to Prevent Relapse of Acute Lymphoblastic Leukemia

Background: The curative potential of allogeneic transplant (alloHCT) in patients with acute lymphoblastic leukemia (ALL) is hampered by relapse, the major cause of treatment failure. Risk factors for relapse include measurable residual disease (MRD) before or after alloHCT, transplantation in second complete remission (CR) or beyond, and reduced intensity conditioning (RIC) regimens. In ALL patients, relapse rates range from 30% to 50%, with most relapses occurring within the first year after alloHCT. After relapse, options for disease control are limited, and consequently overall survival (OS) is poor. Post-alloHCT maintenance may reduce MRD, provide time for a graft-versus-leukemia effect to develop, and ultimately decrease relapse rates and prolong progression-free-survival (PFS) and overall survival (OS). Inotuzumab ozogamicin (INO) is an anti-CD22 monoclonal antibody bound to calicheamicin and approved to treat relapsed ALL. With the hypothesis that low-dose INO would be safe and reduce relapse rates, we conducted a phase I study to define INO dose in this setting. Herein we present the results of this multicenter study. Methods: Eligibility included patients aged 16-75 who underwent alloHCT for CD22+ALL and were in CR after transplant, had a high risk for relapse after alloHCT (MRD before or after alloHCT, transplantation,in second CR or beyond, RIC regimen, lymphoid blast crisis of chronic myeloid leukemia, or Ph-like ALL), had adequate graft and organ function after transplant (platelets > 50 ANC > 1000), did not have active grade III/IV aGVHD or any active liver GVHD, and had no history of hepatic VOD. Patients were consented after transplant. The primary objective was to define a post-alloHCT maximum tolerated dose (MTD) of INO. Secondary objectives included describing the safety profile of INO, determining the rate of hepatic veno-occlusive disease (VOD) in this setting, evaluating non-relapse mortality (NRM), PFS and OS at 1-year post-alloHCT, and determining if INO at these doses is effective at eradicating MRD. Pre-treatment tests included a bone marrow biopsy and aspirate to assess disease state and MRD. The trial design was a 3+3 model. The study treatment consisted of 28-day cycles (initially planned for 4 cycles, increased up to 12 cycles) of INO starting at dose level 0 (0.3mg/m2; n= 4), then 0.4 mg/m2 (n= 3), 0.5 mg/m2 (n= 6) and 0.6 mg/m2 (n= 5) given on cycle day 1. The first cycle was initiated between day 40 and 100 post-alloHCT. Pre-defined dose limiting toxicities (DLT) included VOD, prolonged cytopenia (more than 28 days), severe toxicities ascribed to INO, and death. If there were no DLTs experienced in first two cycles of a cohort, subsequent patients were enrolled in the next highest dose level. Results: We have enrolled 19 patients (18 treated, Table 1) with a median follow up of 18.1 months and a median number of 3 doses (range, 1-9). Only treated patients are included in analyses. The median time to first INO dose was 84 days (range, 47-100) after alloHCT. There was one DLT (prolonged thrombocytopenia) and the study reached the final pre-defined dose of 0.6mg/m2. No VOD was observed. The most common toxicity was thrombocytopenia (42.1% of patients; 31.6% experienced grade 3 or 4 thrombocytopenia), neutropenia (31.6% of patients, 15.8% experienced grade 3 or 4 neutropenia), and nausea/vomiting (42% of patients, all grade 1 or 2). 68% of patients received reduced intensity or non-myeloablative conditioning. OS at 1-year post-alloHCT was 94.4% and PFS was 88.9%. One-year NRM was 5.5%. One patient died of aGVHD on 0.6mg/m2 and one patient relapsed on 0.5mg/m2 at 7 months post-alloHCT. Another patient relapsed from Ph-like ALL 23 months post-alloHCT. Seven of the patients enrolled either had MRD before (n = 4) or after alloHCT (n = 3) as measured by flow cytometry, PCR, or NGS. For patients with MRD before or after alloHCT, 1-year OS and PFS are 100% (Figure 1). The 1-year PFS and OS of recipients of RIC transplants was 83.3% and 91.7%. Conclusions: We have determined that INO at doses of 0.3, 0.4, 0.5 and 0.6mg/m2 was well tolerated and that 0.6 mg/m2 is the MTD. Thrombocytopenia may be the DLT. The low observed relapse rate and favorable safety profile justify investigating low-dose INO as maintenance therapy after alloHCT in a phase II trial. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

The Clinical Observation with Ruxolitinib as Graft-Versus-Host Disease Prophylaxis for Children with Thalassemia after Unrelated or Haploidentical Allo-Hematopoietic Stem Cell Transplantation

To retrospectively analyze the efficacy and safety of ruxolitinib therapy for children with thalassemia after unrelated or haploidentical stem cell transplantation. From March 2020 to March 2021, 22 patients received successfully allogeneic hematopoietic stem cell transplantation in the Zhongshan Hospital of Xiamen University, from +30 to 100 days,those patients received ruxolitinib therapy (2.5 mg, twice daily) and all adverse reactions were observed, include aGVHD, cGVHD, CMV and EBV infection. 22 patients underwent allogeneic stem cell transplantation, 5 patients were diagnosed as aGVHD, 3 patients had grade I－II skin GVHD and 2 patients had grade II intestinal GVHD, those patients were cured. All patients were followed up for more than 21 weeks, 4 cases developed cGVHD, including 3 cases of localized liver GVHD and 1 case of pulmonary GVHD, those were relieved after active treatment. 8 patients had elevated EBV copies (>3×103/ml), and 3 patients had increased CMV copies, the patients recovered after immunosuppressant and antiviral treatment. There was no CMV infection and EBV related post-transplantant lymphoproliferative disorders(PTLD), and no transplant related deaths. Ruxolitinib can effectively reduce the incidence and severity of GVHD without affecting the hematopoietic recovery, and improve the survival status of thalassemia children after transplantation.

A plethora of ocular surface manifestations in a multidisciplinary ocular graft-versus-host disease unit

To describe the experience in a recently created ocular graft-versus-host disease unit in a tertiary hospital and to detail ocular surface features and complications after allogeneic hematopoietic stem cell transplantation (allo-HSCT). This retrospective study included all patients who underwent allo-HSCT, with or without chronic GVHD and were being monitored in the Hematopoietic Stem Cell Transplantation Unit in the UNICAMP Clinical Hospital (Campinas, Sao Paulo, Brazil) from 2015 to 2020. Patients were concomitantly evaluated by hematology and ophthalmology teams of the Ocular GVHD Unit. Hematologists performed a comprehensive systemic evaluation searching and grading mouth, skin, lungs, gastrointestinal tract, liver and genitalia GVHD. While ophthalmologists evaluated ocular symptoms through specific questionnaire (Ocular Surface Disease Index—OSDI) and a protocol of distinct ocular surface parameters for dry eye disease (1) and ocular complications, which encompassed meniscometry, non-invasive tear break-up time (NITBUT) measurement, conjunctival hyperemia quantification, meibography, fluorescein and lissamine staining and Schirmer’s test. Patients were diagnosed with chronic GVHD using the National Institutes of Health (NIH) Consensus Criteria for Chronic Graft-versus-Host Disease. The International Chronic Ocular GVHD Consensus Group (ICOGCG) score was obtained at the onset of ocular disease presentation or afterwards. A total of 82 patients underwent allo-HSCT (97.6% full matched and 2.4% haploidentical), mainly for cases of leukemia and 73.2% had chronic GVHD. Mean onset time for chronic GVHD was 232 ± 7.75 days. The mouth, skin, and eyes were the main organs involved (63%, 50%, and 48%, respectively). Symptom scores and all ocular surface parameters differ in patients with and without chronic GVHD and along different timepoints of the follow-up. Ocular complications mostly involved were severe DED and meibomian gland dysfunction, conjunctival scarring, cataract and infections resulting in keratitis and corneal perforation. As therapeutic strategies, 73% patients received preservative-free lubricants, 27% autologous serum, 48% topical steroids, 27% oral tetracycline derivatives, 22% mucolytic eye drops and 3 patients needed bandage contact lens. Ocular GVHD is a complex and challenging disease with varied manifestations, resulting in a broad range of ocular test endpoints, and inconsistent treatment responses. The main ocular presentations were dry eye, meibomian gland dysfunction and cataracts. The therapeutic approach often involves topical steroids and autologous serum tears. It is important to monitor these patients closely, so the ocular GVHD Unit may improve the care, providing prompt identification of ocular manifestations and faster treatment of complications.

Costs of matched-sibling, unrelated, and haploidentical hematopoietic cell transplantation and risk factors for greater financial burden - a Brazilian FACT-accredited single-center analysis.

The complexity and costs of hematopoietic cell transplantation (HCT) have increased over the last decades with the popularization of unrelated donor (URD) transplantation and the introduction of haploidentical transplantation with posttransplant cyclophosphamide. Few studies have addressed this issue. The objective of this study was to analyze HCT costs in a single FACT-accredited private non-profit hospital. We included 79 patients who underwent HCT between 2018 and 2020. We have included all costs from admission day until D + 180. We used a lognormal regression. Median age was 53 y/o and most donors were unrelated (51%). Costs were higher with haploidentical donor (42%, p = 0.017, compared with URD), higher HCT-CI (15% for each point, p = 0.0056), and in patients with liver or gastrointestinal GVHD (45%, p = 0.033), and lower in patients who received CD34 > 2.5 × 10E6/kg (42%, p = 0.0038). We built a score based on the following risk factors: HCT-CI > 3, CD34 ≤ 2.5 × 10E6/kg, haploidentical donor, and donor age > 30 y/o. Patients with 2 + risk factors (N = 53) had a median cost of USD 226,156.00, compared with USD 93,048.00 in patients with zero or 1 point (N = 26, p < 0.0001). In summary, we have shown that HCT costs are higher with lower doses of CD34 cells, haploidentical HCT (provided that the costs of stem cell procurement and ATG are not included), and in patients with higher HCT-CI. Prospective and refined cost analyses comparing haploidentical and URD transplants, as well as effective strategies for patients with higher HCT-CI scores, are warranted. We found no difference in costs between URD and MSD transplantation.

Use of Belimumab for Prophylaxis of Chronic Graft-Versus-Host Disease

The most transformative approach for controlling chronic Graft-versus-Host Disease (cGvHD) after allogeniec hematopoietic cell transplantation (alloHCT) would be the prevention of its most severe and irreversible clinical manifestations instead of treating already established disease. Belimumab is a monoclonal antibody, approved for treatment of systemic lupus erythematosus and active lupus nephritis, which inhibits binding of B-cell-activating factor (BAFF) to its receptors on B cells, thus inhibiting the survival of autoreactive B cells. Given the role of B cells in cGvHD pathophysiology and the now substantiated role of BAFF in promoting B Cell Receptor signaling in cGvHD (Jia W et al Blood 2021), belimumab might have a role in prevention of cGvHD. We hypothesized that targeting BAFF early after alloHCT would be well-tolerated and have a favorable effect on the incidence and severity of cGvHD.We are presenting data on the first 8/10 patients (Pts) enrolled in the single-center, investigator-initiated phase-1 trial. Belimumab was administered i.v. at 10 mg/kg every 2 weeks for 3 doses followed by 4 doses at monthly intervals, for a total of 7 doses, starting 50-80 days after alloHCT. All Pts were adults who received mobilized peripheral blood grafts from fully matched related or unrelated donors after myeloablative or non-myeloablative conditioning. GvHD prophylaxis was with tacrolimus and methotrexate; one Pt received post-transplant cyclophosphamide. All Pts were in complete remission on day +30 after alloHCT. Pts who received ≥1 dose of belimumab were evaluable for safety and ≥2 doses for efficacy assessment. cGvHD was diagnosed according to the NIH criteria. All analyses are descriptive. Specimens for correlative studies are cryopreserved until the study end.We found that Pts tolerated belimumab well, with no reported grade ≥3 treatment-related adverse events. There were no significant infections or myelosuppression. Seven out of eight Pts on the study received all 7 doses of belimumab, as planned. Of those 7 Pts, 5 Pts are without any evidence of cGvHD and completely off immunosuppression at the median of 18 months (mo) after completing belimumab (median of 24 mo after alloHCT). Seven mo after completing belimumab on study, 1 Pt developed cGvHD involving skin, eyes and liver, and died from complications of pneumonia and liver failure due to longstanding hemochromatosis, iron overload and possible liver GvHD. Another Pt relapsed with leukemia 1 mo after completing belimumab. He is now in remission after treatment with enasidenib and 2 donor lymphocyte infusions (DLI), 12 mo after completing belimumab (18 mo after alloHCT); he developed mild eye GvHD following DLI. One out of eight Pts on the study stopped treatment early, after receiving only 3 doses, due to thrombocytopenia (unrelated to belimumab) and insurance issues. He was found to have relapsed lymphoma 3 mo later, was treated with venetoclax and DLI, and remains in remission with no evidence of cGvHD 15 mo after stopping belimumab. Two additional patients are being enrolled. Immune reconstitution studies are showing that B-cell counts remain low 1 year out of transplant in all Pts (Figure 1).This preliminary data describes for the first time the use of belimumab for prophylaxis of cGvHD. Overall, belimumab was very well tolerated. Only 1 Pt on the study developed cGvHD. It is worth noting that his cGvHD developed 7 mo after completing belimumab suggesting that a longer duration of therapy should be tested in future studies. The 2 Pts with relapsed disease both had high risk malignancies. Risk stratification of high risk patients will be critical in future prospective studies. While more Pts are needed to further assess the impact of prophylactic belimumab on the incidence of cGVHD, these preliminary results are encouraging. Absence of severe infections is reassuring. B-cell reconstitution was delayed, as expected. Ongoing correlative studies will further evaluate BAFF levels and B-cell subset reconstitution after alloHCT. [Display omitted] DisclosuresPusic: Syndax: Other: Advisory Board. Sarantopoulos: Rigel: Other: Advisory Board. Uy: Macrogenics: Research Funding; Jazz: Consultancy; Genentech: Consultancy; Astellas: Honoraria, Speakers Bureau; Novartis: Consultancy; AbbVie: Consultancy; GlaxoSmithKline: Consultancy; Agios: Consultancy. OffLabel Disclosure:Drug: Belimumab This study: Belimumab for prophylaxis of chronic GvHD. Approved indication: Belimumab is approved for treatment of systemic lupus erythematosus and active lupus nephritis.

Chronic liver graft-versus-host disease in allogeneic hematopoietic stem cell transplantation recipients during tapering or after stopping calcineurin inhibitors.

Chronic graft-versus-host disease (cGVHD) of the liver is often observed in allogeneic hematopoietic stem cell transplantation (allo-HSCT) during tapering or after stopping calcineurin inhibitors (CI). We conducted a retrospective analysis of 242 allo-HSCT recipients whose CI dose was reduced to less than 40mg of cyclosporin A or 0.4mg of tacrolimus to clarify the clinical characteristics of liver GVHD in patients on low-dose CI. Sixty patients (25%) developed clinically suspected liver cGVHD while on low-dose CI. Multivariate analysis showed that donor age ≥ 40years [hazard ratio (HR) 2.20], myeloablative conditioning (HR 2.19), female donor to male recipient (HR 2.53) and recipient seropositivity for herpes simplex virus (HR 2.52) were significant risk factors for liver cGVHD during low-dose CI period. Peak aspartate aminotransferase and alanine aminotransferase levels were higher in patients with liver GVHD during low-dose CI period than in other periods. Twenty-seven patients were initially treated with resumption of CI or a CI dose increase and 21 responded. Among the 18 patients treated with corticosteroids, total bilirubin was a risk factor for failure of corticosteroid therapy. The results of this study clarified the clinical characteristics of liver GVHD in patients on low-dose CIs.

Transaminase Elevation As a Marker of Liver GVHD Is Associated with Poor Survival Among Allogeneic Stem Cell Transplant Recipients

Introduction: Liver dysfunction (Hepatotoxicity) is a frequent complication, occurring in 50-72% of patients undergoing allogeneic hematopoietic stem cell transplantation (AHSCT) with mortality rates ranging between 4-15%. Management of liver dysfunction is often complex as multiple etiologies might exist and treatment could be contradicting. A major cause of liver dysfunction in AHSCT is liver GVHD. There is a paucity of data on incidence, risk factors, and outcomes of liver GVHD following AHSCT. In addition, the current liver acute GVHD grading system incorporates only total bilirubin levels. Therefore, a subset of patients with transaminase elevation but normal bilirubin is frequently missed. We aimed to evaluate transaminase elevation as a marker of liver GVHD, confirmed by liver biopsy and/or responsiveness to immunosuppression.Methods: We retrospectively evaluated adult patients who underwent AHSCT between January 2003 and December 2010 at our institution. The primary objective was to compare overall survival (OS) and relapse free survival (RFS) between patients with liver GVHD and other etiologies of transaminase elevation. Additionally, we evaluated frequency and risk factors of liver GVHD in this population.Results: Total 695 patients underwent AHSCT during the study period, and 263 patients developed transaminase elevation. Of these 263 patients, liver GVHD was clinically diagnosed in 46 patients, whereas histologic diagnosis through liver biopsy was made in 86 patients (total 132 patients). Forty-eight patients developed acute GVHD and 84 developed chronic GVHD. The clinical staging of acute liver GVHD is as follows (number of patients): stage 0 (15), stage 1 (7), stage 2 (6), stage 3 (9), stage 4 (11). The histologic gradings are as follows (number of patients): grade 1 (21), grade 2 (46) and grade 3 (19). The 1-year cumulative incidence of liver GVHD in patients with transaminase elevation post-AHSCT was 44.6% (95% CI, 38.4%-50.5%). Of the remaining 131 patients with transaminase elevation, 18% was due to hemochromatosis, 18% to medications 10% to sepsis and 8% to sinusoidal obstruction syndrome (SOS), whereas in 36% of the patients, the cause of the transaminase elevation could not be identified.The median time to development of liver GVHD was 151 (range, 7-1366) days. More patients in liver GVHD group received matched related AHSCT compared to the other group (50% vs 35%). Of 132 liver GVHD patients, 48 patients developed only liver GVHD, 63 patients had overlap with skin GVHD, and 56 patients had overlap with GI GVHD. A proportion of patients (N=15) with transaminase elevation were classified as clinical stage 0 liver GVHD because of absence of hyperbilirubinemia. The AST, ALT, ALP, total bilirubin and ferritin values at the time of liver GVHD onset were 143 (range, 12-1425), 190 (range, 22-1998), 236 (range, 60-1355), 3.8 (range, 0.8-20.1), and 3366 (range, 62-75000), respectively. Transaminases normalized with treatment in 101 patients (77%) with liver GVHD and 86 patients (66%) with other etiologies of transaminase elevation. The median time to LFTs normalization was 35 (range, 0-3706) days for liver GVHD and 45 (range, 2-1616) days for other etiologies of transaminitis. The multivariate analysis demonstrated liver GVHD to be one of the adverse factors for poor OS and RFS (p<0.003). Disease recurrence was the most common cause of death.Conclusion: Liver GVHD is one of the commonly occurring complications, accounting for adverse survival. Therefore, early recognition with immunosuppressive treatment based on clinical suspicion is essential. Current staging system underestimates the true severity of acute liver GVHD because of discrepancies between elevations of transaminases and bilirubin. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Selinexor Maintenance Is Feasible and Tolerable after Allogeneic Stem Cell Transplant (allo-SCT) for Patients with Acute Myeloid Leukemia (AML) and Myelodysplastic Syndrome (MDS)

Background: Selinexor, an exportin 1 (XPO1) inhibitor, has demonstrated anti-leukemic effects as a single agent in relapsed/refractory AML. We hypothesized that selinexor could be used as maintenance therapy after allo-SCT to prevent disease relapse in patients with AML and MDS.Methods: This phase I trial tests selinexor as a maintenance therapy in patients with AML/MDS undergoing allo-SCT (NCT02485535). The primary endpoint was to determine the maximum tolerated dose (MTD) of selinexor after allo-SCT. Patients were enrolled between day 60 to day 100 post allo-SCT if there was no GVHD or disease relapse. Once weekly oral selinexor started approximately 100 days post allo-SCT, and continued for 12 months if tolerated, and there was no relapse of disease. Dose limiting toxicity (DLT) was defined as any grade 3 or higher treatment-related adverse event (AE) that occurred within the first treatment cycle (28 days).Results: From January 2016 to March 2017, nine patients were enrolled including seven with AML and two with MDS. Selinexor dose levels were 60mg weekly (n=6) and 80mg weekly (n=3). Per stem cell transplant disease risk index (DRI), three patients had high risk and five had intermediate risk. Six patients with AML entered allo-SCT in CR (five CR1, one CR2) and one patient had refractory AML. Two MDS patients had active disease at the time of transplant. Four patients had matched related donors (MRD), two had matched unrelated donors (MUD), and three had haplo/cord SCT. One patient had myeloablative conditioning, and eight had RIC. The median time from allo-SCT to first dose of selinexor was 97 days (range: 66-102). One of the first three patients treated at 60mg weekly developed grade 3 nausea, thus three more patients were enrolled to the 60mg weekly dose level. After no additional DLTs at 60mg weekly, patients were enrolled to 80mg weekly dose level. While there was no DLT during the first cycle treatment at 80mg weekly, the dose of selinexor was decreased to 60mg weekly at the beginning of cycle 2 due to decreased chimerism in one patient and in one in cycle 3 due to neutropenia; one patient stopped treatment in the middle of cycle 2 due to leg cramping. The MTD of selinexor post allo-SCT was determined to be 60mg weekly. The most common AEs were fatigue (67%), anorexia (56%), nausea (56%), vomiting (33%) and diarrhea (22%) which were grades 1-2 with the exception of one case of grade 3 nausea. These AEs were manageable with adequate supportive care, including ondansetron, dexamethasone, megestrol and short hold of drug. One patient is finishing 12 months of treatment without major complications. One patient developed grade 1 skin GVHD during tapering immunosuppression not related to selinexor, and 2 patients developed skin and/or liver GVHD after donor lymphocyte infusion (DLI) for low chimerism, which was not related to selinexor. With median follow-up of 359 (219-639) days from transplant, two patients had disease relapse after 2 and 4 cycles of treatment, of which one patient died from disease progression. Three patients are still on treatment, three patients are off study due to tolerability (Neutropenia=1, fatigue=1; and leg cramping=1), and one patient is off study due to liver GVHD after donor lymphocyte infusion (DLI). The median remission duration from the start of Selinexor treatment was 199 (59-542) days. Correlative studies for MRD monitoring and immune reconstitution are on-going.Conclusions: Selinexor maintenance after allo-SCT is feasible and tolerable without increased risk of GVHD. The recommended phase II dose of selinexor maintenance post allo-SCT is 60mg weekly. Additional patients are being enrolled to the expansion cohort at the dose of 60mg weekly for better understanding of the toxicities and efficacy. DisclosuresLiu:BMS: Research Funding; Karyopharm: Research Funding. Larson:Daiichi Sankyo: Consultancy, Research Funding; Amgen Inc.: Research Funding; Novartis: Consultancy, Research Funding; Astellas: Consultancy, Research Funding. Stock:Pfizer: Consultancy, Membership on an entity's Board of Directors or advisory committees; Seattle Genetics: Consultancy, Membership on an entity's Board of Directors or advisory committees; Amgen: Consultancy.

Interim Results of a Phase 2 Clinical Trial Using Mb-IL21 Ex Vivo Expanded NK Cells to Enhance Graft Versus Leukemia Effect for Patients with Myeloid Malignancies after Haploidentical Transplantation

▪Background: Disease relapse has emerged as the most common cause of treatment failure after transplantation. Natural killer (NK) cells have potent antitumor and antiviral effects. We hypothesized that multiple infusions of ex vivo expanded NK cells administered in early post-transplant will enhance graft-versus-leukemia (GVL) effect and began a phase 1/2 clinical trial in haploidentical transplant patients (clinicaltrials.gov NCT01904136).Methods: Results from the phase 1 clinical trial were recently reported. The aim was to determine the maximum tolerated dose of ex vivo expanded NK cells with a K562 expressing mb-IL21 feeder cell system in patients with myeloid malignancies (AML/MDs/CML). Patients received conditioning with melphalan 140mg/m2, fludarabine 160mg/m2 and 2GyTBI, and GVHD prophylaxis with post-transplant cyclophosphamide, tacrolimus and mycophenolate. All patients had bone marrow graft. NK cells were generated from peripheral blood mononuclear cells of the same donor and infused fresh on Day -2, and cryopreserved on Day +7 and +28 (up to 90). Having an NK-cell alloreactive donor or certain KIR genotype was not a requirement to participate in this study, although these characteristics were evaluated in all patients. In the phase 1 study dose escalation patients received NK cells starting at 1x105 to 1x108/Kg/dose for 3 doses. No dose limiting toxicities were observed in the phase 1 study. 1x108/kg/dose was chosen for the phase 2 trial.Results: Twenty patients were treated to date, 13 on the phase 1 study and 7 on the phase 2 part. The median age was 45 years (range 18-59), the median follow-up survivors was 16.5 months. Twelve patients (60%) were females. The median HCT-CI was 2 (range 0-8). Ten patients (50%) had AML (9 in CR1 and 1 beyond CR2), 3 (15%) had MDS (2 with induction failure, 1 untreated), and 7 (35%) had advanced CML (4 in CP2 after progression to blast-phase of which 2 in CNS, 1 in CP1, 1 clonal evolution). Of the patients with AML 4 patients with AML had poor-risk cytogenetics and 6 had intermediate with high-risk mutations except one (2 had FLT3, 2 had NRAS, 1 had ASXL1) and 3 patients had primary induction failure. Two MDS patients had complex cytogenetics and one diploid with high percent of bone marrow blasts. DRI was very high/high in 10 patients and intermediate/low in 10 patients. All patients received all 3 NK cell infusions as planned except one patient. Eight patients received the maximum dose of 1x108/kg/dose.One patient had early death due to hemorrhage. Nineteen patients were evaluable for engraftment and all achieve primary engraftment after a median time of 19 days (range 16-42). Nine of 12 female patients (75%) had donor-specific anti-HLA antibodies. Chimerism was 100% donor in all patients except one who had mixed chimerism. The cumulative incidence (CI) of all aGVHD at Day100 and 1 year post-transplant was 26.1% and 48% (3 patients had late acute GVHD), exclusively grade 2 and exclusively GI tract. No grade 3 or grade 4 aGVHD, and no chronic GVHD was observed. Also, none of the patients developed liver GVHD. The CI of TRM at 1 and 2 years post-transplant was 5.6% and 16.7%, respectively. Only one patient relapsed to date for a CI of relapse at 1 and 2 years on 5.6%. The only patient who relapsed was an advanced FLT3+AML patient (beyond CR2) who received the lowest NK cells dose. None of the patients who received ≥1x105/kg dose relapsed. The median overall survival and progression-free survival (PFS) were not reached. The 1-year and 2-years PFS was 84.4% and 74%, respectively (Figure 1). Eight of 19 patients (42%) reactivated CMV and 4 patients (21%) had grade 1 transient BKV cystitis.Conclusions: Multiple infusions of high doses of donor-derived ex vivo expanded NK cells with a K562 feeder cell system and mbIL-21 administered early post-transplant are safe and are associated with low relapse rate and low incidence of viral reactivation. A randomized trial is needed to confirm these results. [Display omitted] DisclosuresOran:Celgene: Research Funding; AROG: Research Funding; Astex: Research Funding. Lee:Courier Therapeutics, Inc: Membership on an entity's Board of Directors or advisory committees; Cyto-Sen Therapeutics, Inc: Other: Founder, Vice President, Medical Director; Intellia Therapeutics, Inc: Consultancy; Miltenyi: Speakers Bureau.

Impact of Ruxolitinib in Myelofibrosis Allogeneic Stem Cell Transplant Outcome—a Pilot Study

Background: Myelofibrosis (MF) is a clonal stem cell disorder that develops de novo or evolves from a pre-existing myeloproliferative neoplasm such as polycythemia vera or essential thrombocythemia. MF is characterized by dysregulated proliferation of myeloid precursor cells and reactive deposition of connective tissue within the bone marrow, leading to bone marrow fibrosis and eventual failure from ineffective hematopoiesis. Common clinical manifestations include constitutional symptoms such as fatigue, fever, night sweats, weight loss, and hepatosplenomegaly. The clinical course and prognosis of MF is heterogenous, and survival outcomes are largely dependent on risk stratification that incorporates both phenotypic characteristics as well as cytogenetic features. Allogeneic stem cell transplant (alloSCT) is the only available treatment option to cure MF. Ruxolitinib (Rux), a potent JAK1/JAK2 inhibitor, has demonstrated significant effects in improving constitutional symptoms and reducing spleen size in intermediate to high risk patients. Hence, pre-transplant Rux treatment has often been practiced on patients with symptomatic splenomegaly and/or constitutional symptoms. Moreover, there are ongoing studies suggesting improved outcomes in patients treated with Rux before receiving alloSCT. However, the toxicity profile and tolerability profile of Rux treatment in transplant eligible patients remains unclear, especially the impacts of post-transplant Rux maintenance treatment.Method: We retrospectively analyzed the alloSCT outcomes among MF patients who never received Rux, received Rux before alloSCT, and received Rux before and after alloSCT at Penn State Hershey Cancer Institute between 6/2014 and 6/2017.Result: This pilot study analyzed 15 patients with mean age of 58.5 years old (range: 49-70) at the time receiving alloSCT. Ten patients were male and 5 patients were female. Those patients' MF scores were ranging from 2 to 3 (WHO Grade) at diagnosis. Five patients had primary MF and 10 had secondary MF. At diagnosis, all patients had constitutional symptoms, 14 of them had splenomegaly (1 had splenectomy before his MF diagnosis). Seven patients received Rux treatment before alloSCT, 4 received Rux before and after alloSCT, and 4 never received Rux. The mean length of time for Rux treatment before alloSCT was 8.3 months (range:5-20); the mean length of time for receiving Rux post alloSCT was 12.5 months (range:3-23). All patients received reduced intensity conditioning (RIC) regimen (14 patients received Fludarabine/Bu2, 1 patient received Fludarabine/Cyclophosphamide). One patient in the Fludarabine/Bu2 group also received total body irradiation (TBI) as part of conditioning regimen. The mean neutrophils engraft days for patients received Rux before alloSCT was 13.6 (range: 12-19), for patients received Rux before and after alloSCT was 13.3 (range: 12-16), and for patients never received Rux was 16 (range:13-18). All patients were transfusion independent at day 30 (D30) with >95% donor chimerism in peripheral blood. Following alloSCT, 4 patients restarted Rux at a mean time of 3.5 months (range: 3-4.5) post alloSCT. CMV viremia was detected in 5 patients before day 100 of alloSCT; in which 3 patients never received Rux. Fungal pneumonia (1 aspergillus) occurred in 3 patients who received Rux before allSCT. Multiple episodes of GVHD (grade II-III) occurred in 4 patients, however 3 of them never exposed to Rux before. One patient treated with Rux before alloSCT experienced liver GVHD (grade I to II) and responded well to steroids and immunosuppression treatment. The mean spleen size reduction in 6 months after alloSCT among patients received Rux before and after alloSCT was 26% (20 to 32%), among patients received Rux before alloSCT only was 18.2% (6 to 40%), and among patients never received Rux was 17.7% (12 to 24%). All patients are alive and in complete remission.Conclusion:Patients in this pilot study tolerated well to Rux treatment before and after alloSCT with neither prolonged engraft time nor increased toxicity. Before and after alloSCT Rux management may improve overall outcomes. Further study on large patient population is warranted. DisclosuresNo relevant conflicts of interest to declare.

Adverse Effects of Busulfan Plus Cyclophosphamide versus Busulfan Plus Fludarabine as Conditioning Regimens for Allogeneic Bone Marrow Transplantation

Background:The side effects of conditioning regimens on the success rate of allogeneic transplantation around the world have been challenging. In this study, we aimed to investigate the side effect of Bu/Cy and Bu/Flu regimens on our patients who underwent allogeneic bone marrow transplantation. Methods:We analyzed 180 patients receiving bone marrow transplantation in Taleghani Hospital, in Tehran, Iran between April 2016 and December 2019. Patients in group A received a combination of intravenous busulfan 0.8 mg/kg QID over two hours for 4 consecutive days (12.8 mg/kg in total)(Savani et al., 2006) and cyclophosphamide 60 mg/kg per day for two consecutive days. Patients in group B received busulfan the same as the first group in combination with fludarabine equal to 40 mg/m² per day. Patients were followed up at regular intervals up to two years after transplantation. Result:Various items were evaluated for patients, including cardiopulmonary function, psychological disorders, GVHD, and endocrine disorders such as hypothyroidism, fertility, or gonad dysfunction. Primary hypothyroidism developed in 13.3% and 11.1% of the Bu/Cy and Bu/Flu groups, respectively (p=0.230). None of the patients in either group experienced infertility or gonad dysfunction. In group A versus group B, pulmonary diseases were detected in 4.4% versus 6.6% of BMT recipients, respectively (p = 0.223). In both groups, mitral and tricuspid regurgitation were observed in patients (8.9% vs. 11.1%; p = 0.189). Incidence of Psychological disorders was no significant difference between the two groups. 32.2% of group A versus 34.45% of group B had skin and liver GVHD, respectively (p = 0.235). Conclusion:The therapeutic-related adverse effects of the two conditioning regimens in patients who underwent allogeneic bone marrow transplant were almost similar. To improve quality of life and overall survival among BMT patients, careful evaluation of treatment-related complications should be part of the regular follow-up of them.

Prospective analysis of BKV hemorrhagic cystitis in children and adolescents undergoing hematopoietic cell transplantation

BK virus is one of the most common causes of hemorrhagic cystitis (HC) in children undergoing hematopoietic cell transplantation (HCT). Viruses can be found in urine and serum samples of immunocompromised patients. Malignant diseases, age, cell source, day of granulocyte reconstitution, conditioning regimen, or use of total body irradiation may play an important role in BKV epidemiology, development of hemorrhagic cystitis course, and outcome. The aim of this study was to evaluate the incidence, clinical course, and risk factors for BKV-HC in children undergoing HCT. A total number of 133 patients who were prospectively tested for BKV colonization/infection were enrolled into this multicenter analysis. Episodes of BKV-HC occurred in 36/133 (27%) enrolled subjects. In a univariate analysis for BKV-HC incidence, the following factors were significant: age >5 years, peripheral blood transplantation, matched unrelated donor (MUD) transplantation, busulfan-cyclophosphamide-melphalan conditioning regimen, and acute myeloblastic leukemia (AML) diagnosis. Presence of acute graft-versus-host disease (aGVHD) in liver and gut GVHD was a significant risk factor of BKV-HC. No BKV-attributed deaths were reported. In multivariate analysis, the incidence of HC was significantly higher in patients with AML, age >5 years, MUD transplants, and children with GVHD. HC is a frequent complication after HCT among children causes prolonged hospitalization but rarely contributes to death. We identified risk factors of BKV-HC development in children, with focus on aGVHD: we concluded that excessive immune reaction connected with GVHD and immunosuppression drugs might play a pivotal role in the development of BKV-HC.

P2X7 receptor antagonism increases regulatory T cells and reduces clinical and histological graft-versus-host disease in a humanised mouse model.

Graft-versus-host disease (GVHD) is a severe inflammatory response arising from allogeneic haematopoietic stem cell transplantation. Previous studies revealed that antagonism of the P2X7 receptor with Brilliant Blue G (BBG) reduced liver GVHD but did not alter clinical GVHD in a humanised mouse model. Therefore, the present study aimed to trial a modified injection regime using more frequent dosing of BBG to improve outcomes in this model of GVHD. NOD-scid IL2Rγnull (NSG) mice were injected intraperitoneally (i.p.) with 10 × 106 human peripheral blood mononuclear cells (hPBMCs) (day 0), then daily with BBG (50 mg/kg) or saline (days 0-10). BBG significantly reduced clinical score, mortality and histological GVHD compared with saline treatment (endpoint). BBG significantly increased proportions of human regulatory T cells (Tregs) and human B cells and reduced serum human interferon-γ compared with saline treatment prior to development of clinical GVHD (day 21). To confirm the therapeutic benefit of P2X7 antagonism, NSG mice were injected i.p. with 10 × 106 hPBMCs (day 0), then daily with pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid (PPADS) (300 mg/kg) or saline (days 0-10). PPADS increased human Treg proportions compared with saline treatment (day 21), but potential clinical benefits were confounded by increased weight loss with this antagonist. To investigate the role of P2X7 antagonism on Treg survival, hPBMCs were cultured in reduced serum conditions to promote cell death. BBG increased proportions of Tregs (and B cells) compared with saline under these conditions. In conclusion, P2X7 antagonism reduces clinical and histological GVHD in a humanised mouse model corresponding to an increase in human Tregs.

335 - Successful Treatment of Steroid and Ruxolitinib-Refractory Acute Liver GvHD with Single Dose Intravenous Cyclophosphamide

335 - Successful Treatment of Steroid and Ruxolitinib-Refractory Acute Liver GvHD with Single Dose Intravenous Cyclophosphamide