ALBERT

Description: Chronic iron overload associated with hereditary hemochromatosis or repeated red cell transfusions is known to cause cardiac failure. Cardiac arrhythmias have been incidentally noted in patients with iron overload, but often times dismissed as being caused by other co-morbid conditions. Studies with iron-loaded gerbils suggest a role for iron in the development of cardiac arrhythmias, however these studies utilized short duration recordings of anesthetized gerbils. Furthermore, we were unable to reproduce these loading protocols without significant morbidity and mortality. Our goal was to characterize iron-induced arrhythmias in the chronically instrumented, untethered, telemetered gerbil. Monitored gerbils were divided into 2 groups: iron-loaded (n=23) and control (n=8). Iron loaded gerbils received iron dextran intraperitoneally at a dose of 1.7 (n=4), 3.0 (n=5) or 6.2 (n=14) g/kg; control gerbils received dextran. Gerbils were weighed and given a physical exam weekly. Electrocardiograms were recorded for 10 seconds every 30 minutes for approximately 6 months (DSI Ponehma) and reviewed daily. Quantitative analysis was completed on 6 iron loaded (6.2g/kg) and 3 control gerbils. Heart rate and intervals were calculated and arrhythmias were characterized and counted. Cardiac and hepatic histology and tissue iron concentration were assessed. All gerbils showed evidence of frequent sinus arrhythmia (more than one episode per hour). However, except for two control gerbils that showed frequent unifocal PVCs, no significant arrhythmias were noted in daily review. There was no difference in heart rate, P duration, PR interval, QRS duration or QT interval between groups. Neither total number of arrhythmias nor arrhythmias per minute were different between groups. One iron-loaded gerbil had a single episode (11 beats) of supraventricular tachycardia. Two iron-loaded gerbils had PVCs, one had only a single beat and the other had 9 unifocal PVCs over the duration of the study. Iron-loaded gerbils rarely showed other arrhythmias One control gerbil had 260 unifocal PVCs over the duration of the study. Other arrhythmias were noted rarely. Body weight and heart weight was not different between groups, while liver weight increased with increasing iron dose. Cardiac and hepatic iron were significantly increased in iron loaded gerbils when compared to control. Liver weight increased as iron dose increased. Seven of 14 gerbils loaded to 6.2 g/kg developed ascites as assessed both by physical examination and necropsy. We conclude that an iron load sufficient to cause clinical liver disease does not, in the absence of co-morbid conditions, cause cardiac arrhythmias in the gerbil model of iron overload. This suggests that iron alone is insufficient to cause cardiac arrhythmias.

Description: Cardiac events, including heart failure and arrhythmias, are the major cause of death in patients with b-thalassemia. Although cardiac arrhythmias in humans are believed to result from iron overload, excluding confounding factors in the human population is difficult. The purpose of these studies was to evaluate the development of cardiac arrhythmias using the guinea pig model of secondary iron overload. Electrocardiograms were recorded via surgically implanted telemetry devices in guinea pigs loaded intraperitoneally with iron dextran and controls. Cardiac and liver iron concentrations were significantly elevated in the iron loaded animals when compared to control, and were in the range of those reported for humans with thalassemia. Arrhythmias were noted infrequently in both iron loaded and control guinea pigs. No life threatening arrhythmias were detected in either group. These data suggest that iron alone may be insufficient to cause cardiac arrhythmias, and that arrhythmias detected in individuals with iron overload may be the result of a complex interplay of factors.

Description: Abstract 2840 Poster Board II-816 Multiple myeloma (MM) remains incurable with a median survival of 3–4 years. Despite high dose therapy and autologous stem cell transplant (ASCT) most patients relapse with median progression-free survival (PFS) of 2.5–4 years and overall survival (OS) of 4–5 years. Although allogeneic SCT (allo-SCT) is potentially curative due to a graft-versus-myeloma effect, its applicability is significantly limited by high transplant related mortality (TRM). Therefore, the identification of additional independent biological predictors of outcome is required in order to tailor therapy to disease. Natural killer (NK) cells provide first line defence against tumors. NK cells have been shown to recognize and kill myeloma cells both in the allogeneic and autologous settings and donor NK genotype has been shown to influence leukemia free survival following allo-SCT. The aim of this study was to investigate the impact of KIR genotype on event-free (EFS) and OS following ASCT for MM. We performed KIR genotyping on 190 patients with MM receiving a first autologous transplant. KIR genotype and haplotype frequencies were comparable to those published for normal controls. Factors found on univariate analysis to be associated with a shorter EFS included haplotype Bx (containing at least 1 of the KIR B haplotype-defining loci- KIR2DL5, 2DS1, 2DS2, 2DS3, 2DS5, or 3DS1) (median 547 vs 656 days, P = 0.036), ≥3 activating KIR genes (median 547 vs 615 days, P = 0.046), the presence of activating KIR genes KIR2DS1 and KIR3DS1 (median 456 vs 589 days, and 464 vs 619 days, P=0.045 and 0.01 respectively). Disease status at ASCT was the most highly predictive factor for EFS. In patients with good risk disease (CR or PR at ASCT) KIR3DS1 status was highly predictive for EFS 464 days (341–586) vs 731 days (599–862) (P = 0.003) and OS 807days (713-901) vs 967 (925-1009) (P=0.023). KIR3DS1 was not predictive in patients with poor risk disease (P=0.36). Of note KIR3DS1+ve patients were equally represented in good risk (CR and PR) and poor risk (refractory or relapsed) groups at ASCT (around 30% in both groups). Notably the median EFS for KIR3DS1+ good risk patients was not significantly different to poor risk disease patients (P = 0.061). ASCT outcome was then determined according to 3 main groups based on disease status and KIR3DS1 status; A: Good Risk, KIR3DS1-ve; B: Good Risk, KIRDS1+ ve; and C Poor risk (KIR3DS1+ve or -ve). The RR of relapse or death was 1.0, 1.9 (P=0.002, 95% CI 1.3-3.1), and 3.0 (P=0.0001, 95% CI 1.9-4.8) respectively. By multivariate analysis, after adjusting for the presence of adverse cytogenetics and serum albumin and β2m, the KIR3DS1 status and grouping remained highly predictive of poor EFS, RR of 1.0, 2.7 (P= 0.021, 95%CI 1.2-6.2) and 5.3 (P= 〈 0.0001, 95%CI 2.4-11.7) respectively. The prognostic value of KIR3DS1 however, was greatest in patients in whom the ligand for the corresponding inhibitory KIR3DL1, Bw4 was missing. KIR3DS1+ KIR3DL1+ HLA-Bw4 negative patients had significantly reduced median EFS of 400d (315-495) vs 615 (545-684) for all other patients (P=0.048). Again this was most striking in good risk patients. Patients who had the genotype KIR3DS1+ KIR3DL1+ HLA-Bw4 –ve had a significantly shorter EFS survival of 372 days compared to 509 days in KIR3DS1+KIR3DL1+HLA-Bw4+ patients and 793 days for KIR3DS1 negative individuals (P=0.004). In conclusion: Our data from 190 patients with MM suggests that KIR3DS1, a gene previously linked to an increase risk of progression to invasive cervical carcinoma, independently predicts for poor EFS and OS following ASCT. A significant proportion (30%) of patients who are defined as good risk at ASCT (CR and PR) are KIR3DS1+ve and have an EFS which is not significantly different from patients who have refractory/relapsed disease at ASCT. This effect of KIR3DS1 is more significant in the absence of HLA-Bw4, the ligand for the inhibitory receptor KIR3DL1. The mechanism for this is effect is unclear and we are currently performing functional studies to further understand these findings. Disclosures: Apperley: Novartis: Consultancy, Honoraria. Marin:Novartis: Consultancy, Research Funding.

Description: Introduction:Confirmatory laboratory assays for heparin induced thrombocytopenia (HIT) can broadly be classified as functional which have high specificity and rely on activation of platelets by the platelet factor 4 (PF4)-heparin-IgG immune complex or as immune based assays that are relatively more sensitive. At the time when the clinician is evaluating heparin as a cause of thrombocytopenia the 4 "T" criteria are helpful. However, an increase in the patient's platelet count after the heparin has been stopped is critical for confirmation of the diagnosis. Methods: We developed a variation of a previously described technique (Newman Thromb Haemost 1998;80:292) and used clinical criteria as the standard for comparison to evaluate the assay. Radiolabeled 125-I-PF4 is incorporated into the immune complex of PF-4-heparin-immunoglobulin and the amount of radiolabeled immune complex is measured after binding to staphylococcal A protein sepharose (Staph-A). The hospitalized subjects medical record was reviewed to: measure a 4 "T" score, to determine if the patient's platelet count increased after heparin was stopped and to exclude other plausible causes of thrombocytopenia. Aim: The assay relies on the binding of the heparin immune complexes to Staph A. Staph A preferentially binds to larger as compared to smaller immune complexes and the larger complexes produce a greater degree of F(c) mediated platelet activation when compared with the smaller complexes. This suggests the hypothesis that a Staph A based assay will have have better specificity and sensitivity than the currently used methodologies. Results: 28 patient samples were evaluated. True positives were observed in 4 hospital patients and 4 known positives. 19 were true negatives and included 7 from hospital patients and 12 from thrombocytopenic patients who were not treated with heparin. 1 sample was negative in our assay, and was judged as false negative. Concordance between the radiolabeled PF-4 assay and the commercial PF-4 assay was observed in all the 28 patients. Conclusions: To date when judged using clinical criteria, the radiolabeled PF-4 assay correctly distinguished true positives and true negatives in 27 of the 28 samples. Disclosures No relevant conflicts of interest to declare.

Description: Abstract 4361 Introduction: Patients with diabetes mellitus and or metabolic syndrome may or may not be maximally protected from future vascular events with aspirin. To be maximally protected at least 90% of platelets should be inhibited by aspirin (aspirinated). Since platelet acetylation by aspirin is limited to the first 30 minutes after ingestion, in conditions associated with increased platelet production the percentage of aspirinated platelets could be diluted to less than 90% with the usual daily dosing. We studied the pharmacodynamics of aspirin inhibition of platelets to evaluate the feasibility of developing a metric that would detect suboptimal aspirin inhibition of platelets. Methods: The slope of the platelet prostaglandin agonist (PPA) stimulated light aggregometry curve was used to gage the degree of platelet inhibition with aspirin.(Schwartz J. Lab Clin Med 2002; 139:227) Seven normal subjects were studied. After observed ingestion of 325mg of aspirin, the degree of platelet inhibition was studied at the following post aspirin time points; 0, 8hrs, 24hrs and 48hrs. Ex-vivo dilution curves were constructed using time 0 as 100% non-aspirinated and 1hr post aspirin as 100% aspirinated under the assumption that over 24hrs 10% of platelets are newly formed and non-aspirinated. Statistical evaluations were performed for each subject studied using both within subject comparisons and considering the group as a whole, between-subject comparisons. Results: Within group comparisons: A linear regression analysis comparing the in-vivo PPA aggregometry slopes with the PPA slopes obtained via ex-vivo dilutions demonstrated that the ex-vivo dilution curves for a particular subject reliably predicted the degree of aspirin inhibition observed in-vivo for that subject at the following time points with their corresponding percentages of aspirinated platelets; 8hrs (97% aspirinated) 24hrs (90% aspirinated) and 48hrs (80% aspirinated) (r2=0.6, p〈 0.001). In addition, a direct in group comparison showed no differences between the in-vivo PPA slopes and those obtained with ex-vivo dilutions with 8 hr. and 48 hr. time points (p=0.16 and p=0.31, respectively), but a difference was observed at the 24 hr. time point (p=0.02). Entire group comparisons: Platelet inhibition with aspirin was highly variable among subjects. When analyzed as a group, the ex-vivo dilution curves were not predictive of the slopes obtained in-vivo. The PPA slopes observed in-vivo were significantly different from those obtained from the ex-vivo dilution curves at all of the time points (p

Description: Background: IMDs including mucopolysaccharidosis type IH (MPS1/Hurler Syndrome), metachromatic leukodystrophy (MLD), globoid cell leukodystrophy (GLD) and cerebral adrenoleukodystrophy (cALD) are progressive, fatal diseases affecting the central nervous system which are treatable through allogeneic hematopoietic stem cell transplantation (HSCT). CB, in the absence of a matched donor, is the preferred source of stem cells as it is rapidly available and allows greater flexibility in allele matching. As a result of low cell doses, CB transplants in IMD are associated with prolonged periods of neutropenia and reported graft failure rates in up to ~20% (Lum et al 2017 Bone Marrow Transplant 52:846-53; Mallhi et al 2017 BBMT 23:119-25). MGTA-456 is a first-in-class cell therapy produced from a single CB unit using an aryl hydrocarbon receptor antagonist in a 15-day expansion culture of CD34+ cells. In previous phase 1/2 studies, 24 adult and 3 pediatric patients with hematologic malignancies treated with myeloablative conditioning (MAC) and MGTA-456 demonstrated a median 324-fold expansion of CD34+ cells, all patients engrafted, and the time to neutrophil recovery was significantly reduced by a median of 9 days compared to historical controls (Wagner et al 2016 Cell Stem Cell 18:144-55; Wagner et al 2017 Blood 130 supp:662 abstr). Furthermore, higher CD34+ dose has been correlated with improved engraftment and outcomes in IMD transplant patients (Prasad et al 2008 Blood 112:2979-89). Based on these promising data, we postulate the increased CD34+ dose provided by MGTA-456 would reduce the length of neutropenia and risk of graft failure in IMD patients. Patients and Methods: A Phase 2, open-label trial (NCT03406962) initiated in Feb 2018 is enrolling up to ~12 patients

Description: Background Granulocyte colony-stimulating factor (G-CSF) is the standard of care for mobilization of hematopoietic stem cells (HSCs). G-CSF requires 4-7 days of injections and often multiple aphereses to acquire sufficient CD34+ cells for transplant. The number of CD34+ HSCs mobilized can be variable and patients who fail to mobilize enough CD34+ cells are treated with the combination of G-CSF plus plerixafor. G-CSF use is associated with bone pain, nausea, headaches, fatigue, rare episodes of splenic rupture, and is contraindicated for patients with autoimmune and sickle cell disease. MGTA-145 (GroβT) is a CXCR2 agonist. MGTA-145, in combination with plerixafor, a CXCR4 inhibitor, has the potential to rapidly and reliably mobilize robust numbers of HSCs with a single dose and same-day apheresis for transplant that is free from G-CSF. MGTA-145 plus plerixafor work synergistically to rapidly mobilize HSCs in both mice and non-human primates (Hoggatt, Cell 2018; Goncalves, Blood 2018). Based on these data, Magenta initiated a Phase 1 dose-escalating study to evaluate the safety, PK and PD of MGTA-145 as a single agent and in combination with plerixafor. Methods This study consists of four parts. In Part A, healthy volunteers were dosed with MGTA-145 (0.0075 - 0.3 mg/kg) or placebo. In Part B, MGTA-145 dose levels from Part A were selected for use in combination with a clinically approved dose of plerixafor. In Part C, a single dose MGTA-145 plus plerixafor will be administered on day 1 and day 2. In Part D, MGTA-145 plus plerixafor will be administered followed by apheresis. Results MGTA-145 monotherapy was well tolerated in all subjects dosed (Table 1) with no significant adverse events. Some subjects experienced mild (Grade 1) transient lower back pain that dissipated within minutes. In the ongoing study, the combination of MGTA-145 with plerixafor was well tolerated, with some donors experiencing Grade 1 and 2 gastrointestinal adverse events commonly observed with plerixafor alone. Pharmacokinetic (PK) exposure and maximum plasma concentrations increased dose proportionally and were not affected by plerixafor (Fig 1A). Monotherapy of MGTA-145 resulted in an immediate increase in neutrophils (Fig 1B) and release of plasma MMP-9 (Fig 1C). Neutrophil mobilization plateaued within 1-hour post MGTA-145 at doses greater than 0.03 mg/kg. This plateau was followed by a rebound of neutrophil mobilization which correlated with re-expression of CXCR2 and presence of MGTA-145 at pharmacologically active levels. Markers of neutrophil activation were relatively unchanged (20/µl CD34s in 92% (11/12) subjects compared to 50% (2/4) subjects receiving plerixafor alone. Preliminary data show that there was a significant increase in fold change relative to baseline in CD34+ cells (27x vs 13x) and phenotypic CD34+CD90+CD45RA- HSCs (38x vs 22x) mobilized by MGTA-145 with plerixafor. Mobilized CD34+ cells were detectable at 15 minutes with peak mobilization shifted 2 - 4 hours earlier for the combination vs plerixafor alone (4 - 6h vs 8 - 12h). Detailed results of single dose administration of MGTA-145 and plerixafor given on one day as well as also on two sequential days will be presented along with fully characterized graft analysis post apheresis from subjects given MGTA-145 and plerixafor. Conclusions MGTA-145 is safe and well tolerated, as a monotherapy and in combination with plerixafor and induced rapid and robust mobilization of significant numbers of HSCs with a single dose in all subjects to date. Kinetics of CD34+ cell mobilization for the combination was immediate (4x increase vs no change for plerixafor alone @ 15 min) suggesting the mechanism of action of MGTA-145 plus plerixafor is different from plerixafor alone. Preliminary data demonstrate that MGTA-145 when combined with plerixafor results in a significant increase in CD34+ fold change relative to plerixafor alone. Magenta Therapeutics intends to develop MGTA-145 as a first line mobilization product for blood cancers, autoimmune and genetic diseases and plans a Phase 2 study in multiple myeloma and non-Hodgkin lymphoma in 2020. Disclosures DiPersio: Magenta Therapeutics: Equity Ownership; NeoImmune Tech: Research Funding; Cellworks Group, Inc.: Membership on an entity's Board of Directors or advisory committees; Karyopharm Therapeutics: Consultancy; Incyte: Consultancy, Research Funding; RiverVest Venture Partners Arch Oncology: Consultancy, Membership on an entity's Board of Directors or advisory committees; WUGEN: Equity Ownership, Patents & Royalties, Research Funding; Macrogenics: Research Funding, Speakers Bureau; Bioline Rx: Research Funding, Speakers Bureau; Celgene: Consultancy; Amphivena Therapeutics: Consultancy, Research Funding. Hoggatt:Magenta Therapeutics: Consultancy, Equity Ownership, Research Funding. Devine:Kiadis Pharma: Other: Protocol development (via institution); Bristol Myers: Other: Grant for monitoring support & travel support; Magenta Therapeutics: Other: Travel support for advisory board; My employer (National Marrow Donor Program) has equity interest in Magenta. Biernat:Medpace, Inc.: Employment. Howell:Magenta Therapeutics: Employment, Equity Ownership. Schmelmer:Magenta Therapeutics: Employment, Equity Ownership. Neale:Magenta Therapeutics: Employment, Equity Ownership. Boitano:Magenta Therapeutics: Employment, Equity Ownership, Patents & Royalties. Cooke:Magenta Therapeutics: Employment, Equity Ownership, Patents & Royalties. Goncalves:Magenta Therapeutics: Employment, Equity Ownership, Patents & Royalties. Raffel:Magenta Therapeutics: Employment, Equity Ownership. Falahee:Magenta Therapeutics: Employment, Equity Ownership, Patents & Royalties. Morrow:Magenta Therapeutics: Employment, Equity Ownership, Patents & Royalties. Davis:Magenta Therapeutics: Employment, Equity Ownership.

Description: Invariant natural killer T (iNKT) cells are powerful immunomodulatory cells that in mice regulate a variety of immune responses, including acute GVHD (aGVHD). However, their clinical relevance and in particular their role in clinical aGVHD are not known. We studied whether peripheral blood stem cell (PBSC) graft iNKT-cell dose affects on the occurrence of clinically significant grade II-IV aGVHD in patients (n = 57) undergoing sibling, HLA-identical allogeneic HSCT. In multivariate analysis, CD4− iNKT-cell dose was the only graft parameter to predict clinically significant aGVHD. The cumulative incidence of grade II-IV aGVHD in patients receiving CD4− iNKT-cell doses above and below the median were 24.2% and 71.4%, respectively (P = .0008); low CD4− iNKT-cell dose was associated with a relative risk of grade II-IV aGVHD of 4.27 (P = .0023; 95% CI, 1.68-10.85). Consistent with a role of iNKT cells in regulating aGVHD, in mixed lymphocyte reaction assays, CD4− iNKT cells effectively suppressed T-cell proliferation and IFN-γ secretion in a contact-dependent manner. In conclusion, higher doses of CD4− iNKT cells in PBSC grafts are associated with protection from aGVHD. This effect could be harnessed for prevention of aGVHD.

Description: Abstract 1250 Introduction: Patients with occlusive arterial disease are commonly treated with different daily doses of aspirin usually either 81 or 325mg. This treatment practice continues despite studies showing equal protection from future vascular events with daily aspirin doses that exceed 81mg. We evaluated the hypothesis that the amount of platelet inhibition observed 96 hours after initiating aspirin therapy with 3 daily doses of 81mg would be equivalent to that observed using a dose of 325mg. Methods: The amount of platelet inhibition produced when initiating daily aspirin therapy was evaluated using the slope of the platelet prostaglandin agonist (PPA) stimulated light aggregometry curve at 4 time points: time 0 before the first dose, 2 hours after the first dose, 24 hours after the first dose (just before the second dose) and 96 hours after the first dose (24 hours after the 3rd dose). After a 7 day wash out period, during which subjects were instructed not to take aspirin or other NSAIDs, the subjects repeated the study using a 325mg aspirin dose. Because of the large variation in reactivity of platelets among people, each normal subject served as their own control and the results were evaluated using a paired “t” test. We plan to have studied a minimum of 12 normal subjects by the time of the ASH meeting. Results: To date 6 normal subjects were studied. 96 hours after initiating aspirin therapy, 24 hours after the 3rd aspirin dose, there was no difference between the PPA slopes observed for the 81mg=36 and the 325mg=28 (p=.28). Immediate platelet inhibition observed in the 24 hours after initiating aspirin therapy was far greater with the 325mg than the 81mg dose. Two hours after the 81mg dose the PPA slope was 77 compared to the 325mg slope of 12 (p=.000). Similarly, the slopes 24 hours after initiating aspirin were 68 for the 81mg dose and 35 for the 325mg dose (p=.006). The time 0 slopes were also statistically different 81mg=91 and 325=78 (p=.024). Conclusions: 1. Initiation of aspirin therapy with a dose of 325mg dose produces a more immediate inhibition of platelet function with maximal platelet inhibition observed 2 hours after the first dose. 2. Platelet inhibition with an aspirin dose of 81mg occurs more gradually with maximal platelet inhibition observed 24 hours after the 3rd dose at 96 hours. 3. Whether daily aspirin therapy is initiated with a daily dose of 81mg or 325mg, the amount of platelet inhibition observed 96 hours after starting aspirin therapy, 24 hours after the 3rd dose, is equivalent. Comment: This data supports the clinical observations that protection from future vascular events is equivalent with either 81 or 325mg daily dosing and suggests that the current practice of increasing the dose of aspirin in patients who experience a subsequent event may not be beneficial. Disclosures: No relevant conflicts of interest to declare.

Description: Abstract 2539 Invariant NKT (iNKT) cells are a potent, CD1d-restricted immunomodulatory subset of T cells that regulate a variety of experimental immune responses including alloreactivity and acute graft versus host disease (aGVHD).However, their role in human immune responses and in particular in the regulation of clinical aGVHD remains unknown. Given that the frequency of iNKT cells in the blood of normal individuals varies up to 1000-fold we surmised that there was likely to be similar variability in their frequency in peripheral blood stem cell (PBSC) grafts and that the graft iNKT cell dose might be important in the development of aGVHD. To address these hypotheses, using multiparameter flow-cytometry, we determined in G-CSF mobilised PBSC grafts of 61 sibling donors the frequency and the cell dose (in × 106 cells/Kg) of CD3+TCRVbeta11+Valpha24+ iNKT cells as well as of effectors (i.e., CD3+ T cells and CD3-CD56+CD16+ NK cells) or regulators (CD3+CD161+ T cells and CD3+CD4+CD25++FoxP3+ Tregs) of alloreactivity and aGVHD. The median (minimum-maximum) frequency of T cells in the grafts was 62.3% (44.8%–80.7%) of the mononuclear cells and the dose of T cells given with the graft 179.8 (28.7–607.3), while for the NK cells it was 3.3% (0.15%–19.7%) and 9 (0.27–99.6) respectively. The frequency of the CD3+CD161+ cells was 10.5% of the total T cells (1.2%–32.5%) and the dose 17.3 (1.3–120.2). The frequency of Tregs was 5.9% of the CD3+CD4+ population (1.7%–11.9%) and the dose given 6.1 (1.1–36.4). Unlike these cell populations and similar to peripheral blood, iNKT cell content in the grafts varied up to 1000-fold with median frequency 0.045% of the total T cells (0.001%–1.07%) and cell dose 0.075 (0.0014–1.6). There was no correlation between donor age or sex with the frequency or cell dose of any the above cell populations. To assess the role of graft iNKT cells in the development of aGVHD, we selected 41 of the 61 cases, whose G-CSF mobilised PBSC graft was used for a T cell replete, sibling HLA-identical allograft (myeloablative n=34, RIC n=7) for the treatment of haematological malignancies: CML (n=16), AML (n=16), ALL (n=4), MDS (n=3) and myeloma (n=2). aGvHD prophylaxis was cyclosporine-A plus methotrexate (n=39) or cyclosporine-A only (n=2). Only patients who survived 〉100 days were included unless aGVHD developed earlier. Twenty one of the 41 recipients (51%) developed overall grade 0-I aGVHD (group 1) and 20 (49%) grade II-IV aGVHD (group 2). We found no difference in the CD34+ cell dose (median 5.14 vs 5.95, Mann Whitney p = 0.48), sex mismatched allografts (6 cases in each aGVHD group, chi square = 0.92), donor age (42.3yrs vs 46.1yrs, p = 0.4) or recipient age (44.2yrs vs 48yrs, p = 0.4) between the two groups. Similarly, there was no difference in the frequency or cell dose of CD3+ cells (63.3% vs 62.3% p = 0.9 and 179.8 vs 176.5 p = 0.9), CD3+CD161+ cells (14.1% vs 9%, p = 0.17 and 25.8 vs 19.6 p = 0.42), NK cells (3.5% vs 2.9% p = 0.36 and 10.8 vs 9.9 p = 0.35) and Tregs (6.3% vs 5.7% p = 0.68 and 6.1 vs 5.9 p = 0.9). In contrast, there was a significant difference in the graft iNKT cell content between the two aGVHD groups. Specifically, group 1 (grade 0-I) compared to group 2 (grade II-IV) received grafts with an almost 3-fold higher frequency (0.067% vs 0.024% p = 0.026) and dose (0.181 vs 0.07, p = 0.027) of iNKT cells. Further analysis of the relative role of the CD4+ and CD4- iNKT cell subsets revealed that the latter accounted for the protective effect of the high graft iNKT cell content (frequency 0.05% vs 0.014% p = 0.016 and cell dose 0.072 vs 0.023 p = 0.01) but not their CD4+ counterpart (0.038% vs 0.011% p = 0.12 and 0.051 vs 0.029 p = 0.14). Thus, this study is the first to demonstrate that graft iNKT cells are an important determinant of aGVHD in humans and suggests that enrichment of the graft with iNKT cells might be a useful strategy to prevent clinically significant aGVHD. Disclosures: No relevant conflicts of interest to declare.