ALBERT

Description: Abstract 964 Background: High dose immunosuppressive therapy and hematopoietic stem cell transplantation (HSCT) has shown efficacy in severe or rapidly progressive systemic sclerosis (SSc) in phase 1 and 2 trials with durable responses in two thirds of patients (pts), while a recent phase 2 randomised trial in 19 SSc pts showed superior benefit of HSCT over iv pulse cyclophosphamide (Cy) on skin score and lung function. Methods: The ASTIS-trial (Autologous Stem cell Transplantation International Scleroderma trial) is a multinational prospective randomized controlled phase 3 trial, comparing safety and efficacy of HSCT versus Cy in early progressive dcSSc pts with disease duration of a) 4 years or less and evidence of organ involvement or b) of 2 years or less and evidence of systemic inflammation with or without major organ involvement. Major exclusion criteria were: concomitant severe SSc disease (mean PAP 〉 50 mmHg, DLCO 〈 40% predicted, creatinine clearance (CCl) 3months oral) treatment), liver failure. Pts randomized to the transplant arm underwent mobilization with Cy 2×2 g/m2 + G-CSF 10mcg/kg/d, conditioning with Cy 200 mg/kg, rbATG 7.5 mg/kg, followed by reinfusion of CD34+ autologous HSCT. Controls were treated with 12× monthly iv pulse Cy 750 mg/m2. Crossing over was allowed after 2 years. The primary endpoint was event-free survival (EFS), defined as survival until death or development of major organ failure at 2 yrs. Toxicity according to WHO criteria and progression free survival (defined as worsening of modified Rodnan skin score, functional ability, major organ function) were the main secondary endpoints. The effects of treatment were analyzed on an ITT basis and by comparing EFS using the KM survival curves (using log-rank test) and Cox models. Results: 156 pts (female 59%), from 27 centers were enrolled in 10 countries from March 2001 until October 2009 and randomized to HSCT (n=79) or iv pulse Cy (n=77). Seventy-five pts in each arm started treatment, 70 pts in HSCT and 58 pts in control groups completed treatment. The median time (Interquartile Range (IR)) from randomization to completion of treatment was 93 (43.0) days in the HSCT and 338 (41.75) days in the control groups respectively. Baseline characteristics (mean (SD)) of the pts were: age 44 (11.2) yrs, SSc duration 1.4 (1.3) yrs, BMI 24 (14.4), Rodnan skin score 25 (8), HAQ 1.35 (0.8), prior Cy therapy 22%, creat cl 116 (39.5) ml/min, LVEF 65% (8.5), DLCO 59% (14), with no significant differences between the 2 arms. With data cut at 1 May 2012, median follow-up (IR) are 33 (42.0) and 27 (34.0) months in the HSCT and control groups respectively. Forty two events occurred : 18 in the HSCT group (16 deaths and 2 irreversible renal failures) and 24 in the control group (24 deaths). Event-free survival was time-dependent with a hazard ratio at 84 months of 0.22 (95% CI 0.08–0.58, P= 0.002). Eight deaths (including 1 during mobilization and 1 after conditioning) in the HSCT group were deemed treatment-related by the independent data monitoring committee with heart failure (3), ARDS (2), multiple organ failure (2) and pulmonary oedema (1) as the causes of death. In the control group, none died from treatment causes and most deaths were due to progressive disease. Eight pts in the control arm received rescue HSCT treatment, one of whom later died from secondary acute myeloid leukaemia. Two HSCT pts received rescue iv Cy therapy. Conclusions: The ASTIS-trial is the first international, investigator-initiated, phase 3 HSCT trial in early diffuse cutaneous systemic sclerosis. The data show that despite 10% treatment-related mortality, long term event-free survival and overall survival were better in the HSCT group than in the group treated with iv pulse cyclophosphamide. (Funded by the European Group for Blood and Marrow Transplantation, European League Against Rheumatism, AP-HP, NIHR, DIGR, Imtix-Sangstat, Miltenyi-Biotec, Amgen Europe; Current Controlled Trials number, ISRCTN 54371254). Disclosures: No relevant conflicts of interest to declare.

Description: Introduction Since 1996 a joint collaborative effort from the European Group for Blood and Marrow Transplantation (EBMT) and League against Rheumatism (EULAR) collected cases of Autologous Hematopoietic Stem Cell Transplantation (AHSCT) for patients (pts) with severe autoimmune diseases (AD). This retrospective observational study was designed to assess patients outcome and analyse the determinants of treatment responses after AHSCT. Methods All consecutive AD pts treated by AHSCT from 1996 to 2007 according to the EBMT-EULAR consensus statement with ethics committee approved protocol were included. Data were reported yearly via the electronic EBMT data management system PROMISE and outcomes updated as December 2007. Standard AHSCT techniques used either bone marrow (BM), peripheral blood stem cells (PBSC) or both. PBSC were collected with cyclophosphamide CY (CY 4 g/m2) + granulocyte-colony stimulating factor (G-CSF), or with G-CSF alone if cardiac function prevented CY. Cell selection used either CD34+ selection with or without monoclonal antibodies, particularly anti-CD52, anti-CD3, anti- CD19 or anti-CD20. Conditioning regimen used either Total Body Irradiation (TBI) or chemotherapy alone, with various combinations of CY, Busulfan, BEAM ± Antithymocyte Globulins (ATG), which were then subgrouped arbitrarily into high intensity regimen, including Busulfan or TBI, low intensity restricted to CY alone, Melphalan alone and Fludarabine-based regimens, or intermediate regimen with all the other combinations. Primary outcome measure was Progression Free Survival (PFS) defined as survival without evidence of relapse or progression. Secondary outcomes were the Overall Survival (OS) defined as time to death, irrespective of the cause and Transplant Related Mortality (TRM) at day 100, defined as death without ADs relapse or progression. Cumulative incidence curves were used for TRM, considering relapse or progression within 100 days as a competing event and compared using the Gray‘s test. Kaplan-Meier estimate was used to calculate PFS Probabilities and the log-rank test for univariate comparisons. For all pronostic analyses, continuous variables were categorised and the median was used as a cut-off point. Associations of patient, disease and graft characteristics with outcomes were evaluated in multivariate analyses, using Cox proportional hazards for PFS, and proportional hazard regression model of Fine and Gray for NRM. Results: From 1996 to December 2007, 900 AD pts (64 % female, median 35 years) from 171 teams in 27 countries treated by AHSCT were reported to the EBMT data base, mainly multiple sclerosis (MS, n=345), systemic sclerosis (SSc, n=175), systemic lupus erythematosus (SLE, n=85), rheumatoid arthritis (RA, n=89), juvenile idiopathic arthritis (JIA, n=65), vasculitis (VASC, n=26) and hematological immune cytopenia (HIC, n=37). PSC were used as stem cell source for 92% of the pts; 42 % of the pts had in vitro cell selection, mostly with CD34+ selection. All types of conditioning regimens were used in all disease categories with very few patients (7%) with TBI. On the overall population, the 5 years PFS was 43 ± 2 % and OS was 85 ± 1 %. Three years after AHSCT: PFS was 61±5 % for SSc, 55±3 % for MS, 54±6 % for SLE, 52±7 % for JIA, 53±15 % for PMDM, 47±11 % for VASC, 34±9 % for HIC and 23±5 % for RA,); OS was 98 ± 2 % for RA, 93 ± 2 % for MS, 82 ± 5 % for JIA, 83 ± 5 % for SLE and 77 ± 4 % for SSc, depending primarily on AD type (p 35 yrs (HR 0.73, 95%CI (0.59–0.89), p=0.002), AHSCT performed before December 2000 (HR 0.69, 95%CI (0.55–0.85), p=0.008) and number of AHSCT for ADs per centre £ 13 (HR 0.82, 95%CI (0.67–0.99, p=0.05) were associated with a lower PFS. The day 100 TRM was 5 ± 1% overall and appeared significantly lower for a number of patients per centre 〉 13 (HR 0.45, 95% CI (0.21–0.99), p=0.05). ConclusionThe EBMT registry allowed to collect 900 ADs pts with AHSCT after 10 years of collaborative effort. Although retrospective, this largest cohort studied worldwide so far showed that patient age, AD type and number of AHSCT per centre are important determinants of the response. AHSCT appeared a good therapeutic alternative for severe Ads unresponsive to conventional treatment, supporting the ongoing European and north American phase 3 trials comparing AHSCT to standard therapies in SSc, MS, SLE and Crohn’s disease.

Description: Abstract 4537 INTRODUCTION. Autologous HSCT is currently under investigation for the treatment of refractory, progressive MS. A better selection of patients and the use of less aggressive regimens resulted in a significant drop of Transplant-related Mortality (TRM), which is mostly related to immunosuppression. CMV and EBV tend to reactivate in ATG-treated allogeneic HSCT. We investigated the frequency of CMV/EBV viral load in a consecutive cohort of MS patients who underwent HSCT in our Center following BEAM and Anti-T Lymphocyte Globulin (ATG) conditioning regimen. PATIENTS AND METHODS. Thirty-one patients underwent auto-HSCT for severe MS in our Institution from 1999 to 2010. All patients were mobilized by Cyclophosphamide 4 g/sqm + G-CSF and conditioned with BEAM plus rabbit ATG (Thymoglobulin©) at a dose ranging between 7.5 and 10 mg/Kg. Monitoring of CMV/EBV DNA by quantitative PCR on either circulating Mononuclear Cells (MNC) or Whole Blood (WB) was performed in 29 pts. In 18 patients ≥2 tests were available for both CMV and EBV from +20 and +120 days. In 2006, following a case of severe CMV infection, a specific protocol was developed, aimed at monitoring on a weekly basis the viral load in case reactivation until complete negativisation. Pre-emptive treatment was administered in case of viral DNA load 〉1×103 copies/105MNC or 〉1×1044 copies/ml WB in two subsequent determinations. RESULTS. Overall 84 and 95 tests were carried out for CMV and EBV, respectively. 10/29 patients (34.5%) were shown positive for CMV at a median of 33,5 days from HSCT (28-47), whereas 11/29 patients (37.9%) were positive for EBV at a median of 29 days from HSCT (20-109). Five patients resulted positive for both viruses. According to the ongoing protocols, 20% of CMV+ patients and 18% of EBV+ patients received a pre-emptive treatment with a fast drop of viral load. In the other patients the reactivation was self-limiting in 2–3 weeks from the first positive determination. One patient developed a severe CMV infection at + 47 days successfully treated the iv administration of Gancyclovir and Foscarnet. One EBV+ patient developed fever which responded promptly after treatment with rituximab. Moreover, in 16/29 patients a sporadic assessment of CMV/EBV was performed at early (between +5 and + 20) and late phases (beyond 120 days) without any evidence of viral reactivation. CONCLUSIONS. Autologous HSCT with BEAM/ATG can be safely performed in MS patients. A significant reactivation of both CMV and EBV can occur in the 3 months following HSCT, possibly related to the ATG administration. Monitoring of viral load and, whenever necessary, pre-emptive anti-viral treatment should be performed in order to improve safety of HSCT in this subset of non-neoplastic patients. Disclosures: No relevant conflicts of interest to declare.

Description: BACKGROUND. A set of Quality Controls (QC) of a Cord Blood Unit (CBU) at release is required by International Standards for Cord Blood banking, often including Flow Cytometry (FC) counting and viability assessment of thawed stem cells. Reliability and reproducibility of such tests across different CB banks is still an unmet need. Indeed, discrepancies between the thawed product characterized at the Transplant Center and the QC performed at the CB bank, might be addressed to different assessment technologies. Therefore, there is room for a multicenter exercise, aimed to check the reproducibility of the flow cytometry-driven QC and possibly to propose technical standardization and improvements. In this context, a protocol for manipulation and FC assessment of thawed CB samples was circulated and shared among 5 European CB banks(Dusseldorf, Barcelona, Florence, Cremona, Nottingham) , on behalf of Eurocord and Netcord. METHODS. Different techniques of sample preparation were tested on thawed samples, washed samples and the segment attached to the CB bag, respectively, as shown in Fig. 1. A solution of Dextran and Human Albumin as described by the New York CBB was used for dilution/washing the samples. In particular, one sample of each thawed Unit was processed for FC assay undiluted and diluted 1:3 and 1:10, respectively. Five CB Units (CBU) per CB bank unsuitable for banking (with TNC count≥1x109) were selected for the assay. The FC assay protocol was a standard TruCOUNT®, ISHAGE-based (SI), single-platform multigating strategy for CD34+ cells counting, shared by all the Banks in order to standardize the gating strategy. A mock sample was circulated before the exercise in order to check the assay reproducibility. A modification to the ISHAGE methodology (MI) was also proposed with the aim of improving the assessment of CD34+ cells viability in thawed samples. RESULTS A total of 25 CBU across the 5 labs were cryopreserved and thawed following local standards; pre-freezing and post-thawing samples were assessed for CD34+ recovery and viability. Data from all Banks were centralized and statistical analysis was performed. TNC recoveries resulted to be reproducible in all Centers for all the thawed products (undiluted, 1:3 and 1:10 dilution, segment 1:3 diluted and washed undiluted samples, respectively). Both SI and MI strategies showed no significant discrepancies when determining CD34+ absolute number and viability in pre-freezing CBUs (p〉0.05 with pair T test). The recovery of viable CD34+ cells assessed by SI or MI in the thawed products did not show any significant difference in all the experimental conditions, whilst the recovery of total CD34+ cells was always lower in SI-analyzed samples than in MI-analyzed samples (* = pair T test p

Description: Abstract 1221 Poster Board I-243 Introduction A subset of Multiple Sclerosis (MS) patients shows a clinical trend to a fast deterioration of disability despite the use of approved drugs. New immunosuppressive agents are currently employed only in the early phase of the disease but in almost 10% of patients they either do not show any clinical/ radiological improvement or have to be halted for different reasons. Autologous HSCT has been reported as a promising approach for MS patients unresponsive to the available therapies but long term clinical and laboratory follow-up with a stringent MRI monitoring are not yet available. We report here the long term follow-up of a prospective phase II multicenter trial of the Italian GITMO-Neuro cooperative network. Method 21 MS patients (4 Relapsing Remitting and 17 Secondary Progressive) were enrolled between 1999 and 2004 in a prospective trial, aimed to monitor both clinical outcome and MRI imaging. Patients were shown to be refractory to conventional treatments, had a EDSS between 5.0 and 6.5 and at least 1 Gadolinium (Gd) enhancing area in brain MRI. PBSC were mobilized with Cyclophosphamide (4g/m2 ) and Filgrastim; patients were conditioned with BEAM plus rabbit ATG (Thymoglobulin®, Genzyme) and infused with unmanipulated graft. The effect of HSCT was evaluated with serial monthly Gd-enhanced brain MRI for a pretreatment period of 3 months and compared with serial monthly Gd-enhanced MRI imaging for the following 6 months. Subsequently, MRI scans were carried out at, +9, +12, +18 + 24 and at the last follow-up. Clinical outcome was evaluated by both EDSS assessment and number of clinical relapses after the transplant. The same MRI scanning protocol was used at each neuroradiological examination: T1 and T2 total lesion load, new T2 lesions, new hypointense lesions, Gd enhancing activity and progression of brain atrophy were evaluated. Results All patients showed a sustained engraftment with modest early side effects, as previously reported (Saccardi R. et al. Blood 2005 105:2601-07). At a median follow up of 7 years (range 4-10 years) no late effects were reported. Two patients had a clinical relapse 5 years after the HSCT, spontaneously recovered. Nine patients showed a clinical progression at a median of 30 months (16 – 66) after transplant, while the others are either stable (7) or improved (5), as compared with baseline. No patients received any immunosuppressive treatment after HSCT. At MRI examination total lesion load assessed by T2 and T1 sequences remained stable through the follow-up, and only 5 new T2 lesions appeared after HSCT. No Gd enhancing activity was ever observed after transplantation in the examined cases. Brain atrophy progression was higher in the first 2 years after transplantation and then decreased significantly in the following years Conclusions The clinical and MRI results of this prospective study are extremely positive, considering that the majority of cases remained stable at a median of 7 years after transplantation. MRI activity, as evaluated with Gd enhancing areas and the appearance of new lesions, was absent or negligible. Also brain atrophy, rapidly progressing early after the transplant probably due to the suppression of the inflammation, was thereafter shown to be in the range of MS patients. The duration of the follow-up and the stringent MRI methodology provide an evidence of the efficacy of this procedure in this subset of aggressive MS patients. Disclosures No relevant conflicts of interest to declare.

Description: Background: The use of mismatched unrelated (MMUD) donor grafts in leukemic patients who are in need of allogeneic hematopoietic cell transplantation (HCT) and do not have a fully matched donor, represents an alternative therapeutic option. Data on HCT outcome after ≤6/8 MMUD are limited. Methods: We used the data set of the acute leukemia working party (ALWP) of the European society of blood and marrow transplantation (EBMT) in order to assess the transplantation outcome in patients (pts) with acute myelogenous or lymphoblastic leukemia (AML/ALL) transplanted from ≤6/8 MMUD (2-4 HLA mismatches at the allelic level at loci A, B, C and DR) in remission (CR), with no ex vivo T cell depletion (TCD). HCT were performed between 2000 and 2017. Multivariate analyses (MVA) were performed using the Cox proportional hazard model. Results: The study population consisted of 465 patients who met the inclusion criteria (AML 320, ALL 145). The median age at transplantation was 50.2 years (range, 18-71.8) and 34.8 years (range, 18.2-62.9); 54% and 64% were males in AML and ALL groups, respectively. Median follow up was 63 and 75 months, respectively. 69% and 66% were in CR1, while 31% and 34% were in CR2. 40% of AML pts had intermediate risk cytogenetics, 10% poor risk and 5% good risk (missing -45%), respectively. 37% of ALL pts were Ph positive, 22% Ph negative and 18% T ALL (17% B ALL with no Ph data, 6% missing immunophenotyping data). The number of HLA mismatches (HLAMM), it was 6/8 HLA MM - in 82% and 85%, 5/8- in 11.5% and 12% and 4/8- in 6% and 3% of AML and ALL pts, respectively. 85% of AML pts and 77% of ALL pts received PBSC graft. The conditioning regimen was myeloablative in the majority of patients (89% of AML and 61% of ALL pts). 82% and 77% underwent in vivo TCD, respectively. Graft versus host disease (GVHD) prophylaxis was CSA/MTX in 51% and 67% and CSA/MMF in 21% and 10% of AML and ALL pts, respectively. Engraftment was achieved in 97% and 95% of pts, respectively. The incidence of acute (a) GVHD grade (Gr) II-IV and Gr III-IV was 34% and 41% and 14% and 21% for AML and ALL pts, respectively. Total and extensive chronic(c) GVHD rates at 2y were 38% and 38% and 23% and 14%, respectively. The main causes of death were disease recurrence (37% in both), GVHD (29% and 22%) and infections (22% and 26%). In the entire population, 2 and 5y outcomes were: relapse incidence (RI) -28% (95% CI: 24 - 32) and 33% (95% CI: 28 - 37), non-relapse mortality (NRM)- 28% (95% CI: 24 - 32) and 30% (95% CI: 26 - 35); leukemia free survival (LFS) -44 % (95% CI: 39 - 48) and 37 % (95% CI: 32 - 41 ), overall survival -50% (95% CI: 45 - 55 ) and 41% (95% CI: 36 - 46) and GVHD free relapse free survival (GRFS)- 33% (95% CI: 29 - 38) and 27% (95% CI: 22 - 31), respectively. More than 2 HLA mismatches were associated with a higher incidence of aGVHD Gr II-IV (29% vs 15%, respectively). In MVA, RI was lower for ALL vs AML HR (95% CI) 0.60 (0.39-0.93), p=0.02. Disease status (CR2 vs CR1) was poor prognostic factor for RI, LFS and OS. Age was prognostic factor for NRM, LFS and OS. HLA- mismatch at locus DR was poor prognostic factor, giving increased NRM: HR (95% CI) 1.68 (1.1-2.5), p=0.02, and lower LFS: HR (95% CI), 1.42 (1.03-1.95), p= 0.03 and OS HR (95% CI) 1.46 (1.04-2.03), p=0.03 and higher GVHD Gr II-IV HR (95% CI) 1.48 (1-2.17), p= 0.048. Results of MVA in AML pts in CR1 were similar. GVHD prophylaxis with CSA/MMF in comparison to CSA/MTX resulted in higher NRM HR (95% CI) 2.2 (1.27-3.82), p=0.005 and lower GRFS HR (95% CI) 1.58 (1.06-2.35), p= 0.02. Notably, in 154 pts with AML in CR1 transplanted from 6/8 UD with in vivo TCD, C mismatches was associated with higher incidence of cGVHD, HR (95% CI) 2.52 (1.40-4.53), p= 0.002. Conclusions: HCT from ≤6/8 MMUD is an alternative transplantation option for acute leukemia pts in CR that can provide 40% 5y OS, 37% LFS and GRFS of 27%. DR mismatch is a poor prognostic factor. cGVHD incidence at 5y was 41% with 21% being severe and is associated in AML pts with HLA C mismatches. CSA and MTX is the preferred GVHD prophylaxis. Emerging novel agents will hopefully reduce the incidence of cGVHD in the near future and thus improve HCT from ≤6/8 MMUD outcome. Disclosures Rambaldi: Italfarmaco: Membership on an entity's Board of Directors or advisory committees, Other: travel support, Research Funding, Speakers Bureau; Omeros: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Gilead: Membership on an entity's Board of Directors or advisory committees, Other: travel support, Speakers Bureau; Celgene: Membership on an entity's Board of Directors or advisory committees, Other: travel support, Speakers Bureau; Roche: Membership on an entity's Board of Directors or advisory committees, Other: travel support, Research Funding, Speakers Bureau; Amgen: Membership on an entity's Board of Directors or advisory committees, Other: travel support, Research Funding, Speakers Bureau; Pfizer: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Novartis: Membership on an entity's Board of Directors or advisory committees, Other: travel support, Speakers Bureau; Jazz: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau. Socie:Alexion: Consultancy. Mielke:Celgene: Honoraria, Other: Travel support (via institution), Speakers Bureau; EBMT/EHA: Other: Travel support; Bellicum: Consultancy, Honoraria, Other: Travel (via institution); IACH: Other: Travel support; Jazz Pharma: Honoraria, Other: Travel support, Speakers Bureau; DGHO: Other: Travel support; Kiadis Pharma: Consultancy, Honoraria, Other: Travel support (via institution), Speakers Bureau; GILEAD: Consultancy, Honoraria, Other: travel (via institution), Speakers Bureau; ISCT: Other: Travel support; Miltenyi: Consultancy, Honoraria, Other: Travel and speakers fee (via institution), Speakers Bureau. Platzbecker:Celgene: Consultancy, Honoraria; Abbvie: Consultancy, Honoraria; Novartis: Consultancy, Honoraria. Finke:Riemser: Honoraria, Other: research support, Speakers Bureau; Neovii: Honoraria, Other: research support, Speakers Bureau; Medac: Honoraria, Other: research support, Speakers Bureau. Mohty:Jazz Pharmaceuticals: Honoraria, Research Funding.

Description: Background: While intensive consolidation therapy with autologous stem cell transplantation (ASCT) can secure a remission in selected Acute Myeloid Leukemia (AML) patients with intermediate-risk cytogenetics, a substantial proportion will ultimately relapse. Knowledge of the mutational status of FMS like tyrosine kinase 3 internal tandem duplication (FLT3-ITD) and nucleophosmin (NPM) 1 and their possible combinations could further refine a subset of intermediate cytogenetic risk patients who may benefit from ASCT. However, data are limited. We, therefore, set out to evaluate the impact of FLT3-ITD and NPM1 in a large cohort of patients undergoing ASCT. Methods: This was a retrospective analysis of the Acute Leukemia Working Parity of the European Society for Blood and Marrow Transplantation (EBMT) registry. We included 405 de-novo AML patients, from 51 European centers, with intermediate-risk cytogenetics (Grimwade et al., Blood 2010) and complete data on FLT3-ITD and NPM1 status, receiving an ASCT at first complete remission (CR1), between 2000-2014. Leukemia free survival (LFS) was the primary outcome. Secondary outcomes were overall survival (OS), transplantation-related mortality (TRM), and relapse incidence (RI). The latter two were considered as competing events. Univariate and multivariable Cox regression models, adjusted for recipient sex, age, Karnofsky performance status, FLT3/NPM1 combinations, days from diagnosis to transplantation, stem cell source -bone marrow or peripheral blood (PB), and use of total-body irradiation-based conditioning. Results: Patients included had a median age of 52 years and received an autograft at median of 5 months from diagnosis. PB-based autograft and non-TBI conditioning were used in the majority of patients (93% and 90%, respectively). FLT3-/NPM1- was the leading molecular combination (50%), followed by FLT3-/NPM1+ (30%), FLT3+/NPM1+ (11%), and FLT3+/NPM1- (9%). Age, time from diagnosis to transplantation, graft source, and use of TBI were similar between the molecular subgroups. The median year of transplantation was earlier in NPM1- patients (FLT3+/NPM1- 2008, FLT3-/NPM1- 2009, FLT3-/NPM1+ 2010, FLT3+/NPM1+ 2011, p

Description: Abstract 3164 Background. Cryopreservation of Hematopoietic Stem Cells (HSC) is a standard procedure for both allogeneic cord blood and autologous HSC transplantation. PBSC are routinely used for the latter; the graft is usually thawed bedside and shortly reinfused without any manipulation. Infusion-related toxicity is usually mild, although severe reactions have been reported, mostly in patients with co-morbidities at baseline. Indeed, cell viability decreases over time after thawing. Washing the thawed graft was reported to improve both clinical tolerance and cell viability. It is however not routinely carried out as it is time-consuming and may result in clinically relevant HSC loss. The fully automated solution SmartWash™ (Biosafe, Switzerland) is designed to wash a thawed graft in a clinical setting. We report here its validation through an international, prospective, multicenter trial. Methods. A pre-clinical and a clinical study were designed and sequentially carried out in 4 European Centers (Basel, Florence, Marseille and Murcia) to assess the reproducibility and feasibility of the SmartWash™ system. During the pre-clinical phase, 10 PBSC Units/Center (total: 40 procedures) from patients who permanently lost the indication to the transplant were thawed and split in two aliquots: one was washed and the other was left unmanipulated. A shared protocol targeted to assess recovery, viability and stability of thawed PBSC at different time-points was developed; in particular, an ISHAGE-modified flow cytometry method was implemented in order to improve the evaluation of thawed CD34+ cells. Clinical grade hydroxyethyl starch (Voluven® 6%,Fresenius, Germany) was validated and used as washing solution. Following the validation of the method, a prospective, observational clinical trial was started after approval of the competent Authorities. Twenty-five consecutive patients across the 4 Centers underwent an autologous PBSC transplantation according to the local practice and indications; all the grafts were washed by Smartwash™. Cellular recovery, clinical tolerance and transplant outcome were collected according to a shared protocol. Results. All the washing procedures were completed without any laboratory problems in the participating centers. In the pre-clinical study the recovery of total CD34+ cells in the washed product was 101.2%±27.0 as compared to the pre-freezing value, with a viability of 87.2%±10.0, always showing an advantage over the unwashed samples at all considered time-points (0,1,4 and 24 hours after washing, respectively, Fig. 1). The workload of the procedure, as compared to the unwashed process, resulted in a 40 minutes prolongation for each thawed product, among which only 12 minutes were devoted to active time work, whilst 28 minutes consisted in automatic processing. In the clinical trial, patients received a median of 4.32×106 CD34+ cells/Kg (1.32–27.92), as measured before cryopreservation. Recoveries of both total and viable CD34+ cells after thawing and washing are reported in the Fig. 2, replicating the results of the pre-clinical study. Infusion-related toxicity was negligible (1 grade-1 toxicity out of 25 patients). All patients achieved both neutrophil and platelet sustained engraftment at a median of 11 (9–19) and 13 (7–19) days, respectively. Conclusions. SmartWash™ is an efficient and safe automated method to wash thawed PBSC grafts in a closed, clinical-grade system. Its feasibility and reproducibility was confirmed in an international, multicenter, clinical study. The washed product is stable and clinically feasible for several hours after thawing. Disclosures: Saccardi: Biosafe SA: Research Funding. Boieri:Biosafe SA: Research Funding. Loutan:Biosafe SA: Employment.

Description: Introduction: Donor and recipient polymorphisms of immune defense and inflammatory cytokine genes have been associated with outcomes after bone marrow or peripheral blood stem cell transplantation (SCT), but have not been described in the setting of umbilical cord blood transplantation (CBT). Material and Methods: We evaluated the impact of genetic polymorphisms of recipients and CBU units on survival and other CBT outcomes using the following candidate genes related to immune response: NACHT-leucine-rich repeat with a pyrin domain at N-terminus (NLRP1-rs5862, NLRP2-rs043684, NLRP3-rs10754558), Toll-interleukin-1 receptor domain containing adaptor protein (TIRAP/Mal-rs8177374), interleukin-10 (IL10-rs1800872), V-rel reticuloendotheliosis viral oncogene homolog (REL-rs13031237), tumor necrosis factor receptor superfamily - member 1B (TNFRSF1B-rs1061622) and associated protein 4 of cytotoxic T lymphocyte (CTLA4-rs3087243). Seven Netcord banks provided pre-transplant DNA samples from 851 CBU units and 173 recipients. All patients underwent CBT at EBMT centers. Malignant (n=696) and non-malignant (n=155) diseases were analyzed separately. Results: Among the 851 recipients, 57% were male and 61% adults. Degree of HLA matching between CBU unit and recipient (HLA-A, -B at antigen level and -DRB1 at allele level) was 6/6 in 12%, 5/6 in 40%, and ≤4/6 in 48% of transplants. Myeloablative conditioning was used in 77% of cases and anti-thymocyte globulin or monoclonal antibodies in 82% of cases. Median infused total nucleated cells (TNC) and CD34+ cells were 3.7x107/kg and 1.6x105/kg, respectively. In the cohort with malignant diseases, multivariable analysis adjusted for patient-, donor- and disease-related variables showed that recipients of CBU with GG CTLA4 genotype had a poorer neutrophil engraftment (HR 1.24; 95% CI 1.09-1.40; p=0.03), increased NRM (HR 1.52; 95% CI 1.35-1.72%; p