Post Allogeneic SCT Research Articles

CD123 As a Therapeutic Target in Accelerated and Blast Phase Myeloproliferative Neoplasms (MPNs)

Background: CD123 expression in MPNs has been reported in case series and CD123 has been a therapeutic target in one phase I study of patients with chronic myelomonocytic leukemia or myelofibrosis in chronic phase. CD123 has also been described as a marker of leukemic stem cells and incorporated in chimeric antigen receptor T-cell products for treatment of acute myeloid leukemia in clinical trials. Herein, we describe the clinical phenotype of patients with accelerated (AP) or blast phase (BP) MPNs paired with CD123 co-expression on CD45(dim) myeloid blasts. Methods: This is a retrospective, single-institution study of patients with AP and BP MPNs. Subjects were identified by search for MPN diagnoses in the Ohio State University Comprehensive Cancer Center Leukemia Tissue Bank (OSUCCC-LTB). Relevant clinical data was collected by retrospective chart review with approval of Institutional Review Board. CD123 expression was measured by immunophenotyping of cryopreserved peripheral blood (PB) or marrow (BM) samples. Blasts were identified by CD45(dim) gating of lymphocytes. Lymphocytes were also gated on CD7, CD13, CD33, CD34, and CD117. CD123 expression was stratified by quartile and correlation with disease phase, symptoms, splenomegaly, transfusion-dependence, and molecular mutations measured by Chi-square test. Results: Forty-one PB or BM samples from 24 subjects were identified from the OSUCCC-LTB. Nine subjects had paired PB and BM specimens and 6 subjects had samples obtained at multiple timepoints in the course of their disease. Median age at time of sample collection was 65.8 years (range 35.2-79.7) and median time from initial diagnosis to sampling was 13.1 months (range 0-96.5). Constitutional symptoms were present at the time of collection in 24 specimens (58.5%) and splenomegaly present in 20 specimens (51.3%). At sample collection, 19.5% had never received treatment, 19.5% had been previously treated, and 61% were currently on treatment. Most patients on treatment (n=16, 64%) were receiving JAK inhibitor at sample collection. Nineteen samples were obtained in chronic phase (46.3%), nine in AP (22.0%), and thirteen in BP (31.7%). Median WBC count was 4.37K/uL (range 0.3-174.6), median peripheral blasts were 3.2% (range 0-76%), median hemoglobin was 8.8 g/dL (range 5.6-13.2), and median platelets were 42K/uL (range 5-564). Sixteen samples (39.0%) were from transfusion-dependent patients, of which, 68.8% were both red blood cell (RBC) and platelet transfusion-dependent, 25.0% were RBC transfusion-dependent only, and 6.2% were platelet transfusion-dependent only. Median lactate dehydrogenase at time of sampling was 300 U/L (range 116-2,142). Median bone marrow blasts at time of sampling were 3% (range 0-82%). Molecular sequencing and karyotype +/- 30 days of sample collection were available for 18 (43.9%) and 27 (65.9%) of specimens, respectively. The most common mutations identified were JAK2 (50%), ASXL1 (27.8%), SRSF2 (22.2%), CALR (16.7%), and TP53 (16.7%). Normal karyotype was reported in 37% of the samples (one was consistent with donor chimerism post-allogeneic SCT), 33.3% had complex karyotype, 11.1% had del 5q or 7q/-7, and 18.5% had del 13q or 20q. CD123 expression on the CD45(dim) gated blasts ranged from 0.3-95.6% with a median value of 11.02%. CD123 expression is presented in figure 1 box plot. Figure 2a shows correlation between CD123 expression and AP/BP compared to chronic phase disease (p < 0.01). Figure 2b shows CD123 expression correlation with ASXL1 compared to unmutated ASXL1 (p =0.023). There were no statistically significant associations between CD123 expression and any other tested variables. Conclusions: These single-institution data suggest that high CD123 expression on blasts is associated with leukemic transformation and high-risk disease. This is demonstrated by correlation of higher expression in AP and BP MPN as well as correlation with ASXL1 mutation. CD123 expression in MPNs has been previously reported and explored as a therapeutic target in a single phase I trial, though these data suggest that specifically targeting high risk disease, defined by blasts >10% or the presence of ASXL1 mutation, may be more effective with greater target availability. This is an especially important subpopulation of patients with MPNs in need of more effective and less toxic therapy than current standards of care that typically include cytotoxic chemotherapy.

Genomic analysis for TIM-3-expressing measurable residual disease post allogeneic SCT

Genomic analysis for TIM-3-expressing measurable residual disease post allogeneic SCT

AML-142 Patients at High Risk of Relapse Post-Transplant: A Phase 1 Study Design With a Novel Treatment Strategy Using the Estimand Framework

The risk of disease relapse in patients allografted for acute myeloid leukemia (AML) is high and remains the main cause of transplant failure. A potent graft-versus-leukemia (GvL) effect is exerted post-stem cell transplantation (SCT); strategies that may augment a GvL effect have the potential to reduce relapse risk post SCT. Siremadlin/HDM201 is a novel investigational MDM2 inhibitor with single-agent (SA) anti-AML activity. MDM2 inhibitors possess potent immunomodulatory effects (IME) in murine solid tumor and AML models. The post-allogeneic SCT setting is ideal for investigating IME of MDM2 inhibition on the GvL effect. To present a study design assessing the safety and preliminary efficacy of treatment with monotherapy siremadlin and in combination with donor lymphocyte infusions (DLIs) for AML post SCT using the estimand framework (ICH E9 [R1] addendum). Phase Ib/II proof-of-concept. AML patients in CR post SCT at high risk for relapse based on pre-SCT risk factors. The estimand framework mandates definition of experimental intervention according to the clinical question. Due to the risk of GvHD with DLI early post SCT, siremadlin will first be given to eligible patients as priming monotherapy to prevent early relapse; patients who tolerate siremadlin with no evidence of GvHD are eligible to start siremadlin/DLI. Post combination, patients may continue maintenance SA siremadlin, which intends to prolong/enhance the GvL reaction to eradicate residual leukemic blasts. According to their treatment journey, patients may receive only part of the intervention's sequence (e.g., siremadlin priming monotherapy patients not eligible for DLI combination will continue priming). To determine 1) the siremadlin recommended dose (RD) across all parts of the treatment strategy and 2) preliminary efficacy regardless of the treatment parts received. Siremadlin RD will be determined using a Bayesian logistic regression model to guide dose finding of siremadlin during priming/Cycle 1 and a Bayesian time-to-DLT model to assess the siremadlin combination dose. Efficacy will be assessed by the proportion of patients remaining in CR ≥6 months. The estimand framework guided the design of an AML clinical study post SCT to conceptualize the assessment of safety and efficacy of the treatment strategy, accounting for different patient journeys.

Fluconazole vs. voriconazole prophylaxis for prevention of invasive fungal infections post haploidentical stem cell transplant.

e19504 Background: IFIs are one of the most detrimental complications of profound and prolonged neutropenia post-allogeneic SCT. Prophylaxis with broad spectrum azole antifungals has become standard of care given favorable outcomes associated with their use in this setting. Nonetheless, the choice of antifungal agent varies between institutions due to absence of consensus guidelines in patients undergoing haploidentical SCT. Methods: Non-interventional retrospective study seeking to compare efficacy of fluconazole vs. voriconazole for prevention of IFI post haploidentical SCT for hematologic malignancies. SCT databases at two academic insitutions (one uses fluconazole and other uses voriconazole) were queried to identify patients who underwent allogeneic SCT from April 2012 until March 2018, and received prophlaxis with either voriconazole (group I) or fluconazole (group II). Patients with intent to start prophylaxis with fluconazole or voriconazole were included, even if antifungal agent was changed later. Primary endpoint was cumulative incidence of breakthrough IFIs (based on MSG criteria) on day +100 and +180. Fisher exact test was used to compare rates of breakthrough IFIs between two cohorts. Secondary endpoint was overal survival (OS). Kaplan Meiers analysis was performed to compare OS on day +180 and 1-year post SCT. Results: The cohort included 141 patients (group I: 75 and group II: 66). Percentage of patients who underwent transplant for leukemia was higher in group I compared to group II (77.3% vs. 56.1%, p < 0.001). A haploidentical SCT was performed in 62.7% of patients in group I compared to 100% in group II (p < 0.001). All patients received non-myeloablative conditioning. There was no statistically significant difference in rates of IFIs between two groups on day +100 (0% in I vs. 4.5% in II, p = 0.9) and +180 (1.4% vs. 4.6%, p = 0.2). A significant difference in OS was noted on day +180 and at 1-year in univariate analysis (Table). Conclusions: Our findings suggest that prophylactic use of either voriconazole or fluconazole after haploidentical SCT is associated with low rates of IFIs. However, survival data favored voriconazole over fluconazole. [Table: see text]

Therapy of Paediatric B-ALL with a Fast Off Rate CD19 CAR Leads to Enhanced Expansion and Prolonged CAR T Cell Persistence in Patients with Low Bone Marrow Tumour Burden, and Is Associated with a Favourable Toxicity Profile

Introduction: The CARPALL study (NCT02443831) employed a novel CD19CAR (CAT-41BBz CAR) with a faster off rate than the Kymriah FMC63-41BBz CAR (CAT 3.1x10-3s-1, FMC 6.8 x 10-5s-1), with equivalent on-rate (CAT 2.2 x 105, FMC 2.1 x 105). We herein report updated outcomes and CAR T cell persistence with an additional 6 months follow up from a submitted manuscript (Ghorashian et al., Nat Med, submitted) Methods: Patients aged <25 years with high risk, relapsed CD19+ B-ALL were eligible on this multi-centre, open label, non-randomised phase I study of autologous CAT-41BBz CAR T cells. Patients were followed to a data cut-off of 07/18/2019. CAT-41BBz CAR T cells were generated by magnetic bead activation of leucapheresed PBMCs, lentiviral transduction, followed by bioreactor expansion and magnetic bead removal prior to cryopreservation. All patients received lymphodepletion (fludarabine + cyclophosphamide) followed by 1x106/kg CAR T cells. Presence of CAR T cells in the blood and bone marrow (BM) was assessed (flow cytometry and qPCR) monthly for 6 months, then 6 weekly to 1 year and then 3 monthly. BM MRD was assessed (IgH qPCR, flow cytometry) at the same time-points up to 2 years to establish durability of responses as a stand-alone therapy. Primary end-points were incidence of grade 3-5 toxicity and the proportion of patients achieving molecular remission. Results: Of 17 patients recruited, 14 were treated due to manufacturing failure in 3 patients.The median age was 9 years (range 1-19 years). All patients had advanced ALL with a median of 4 prior therapy lines. 10 of 14 patients (71%) had relapsed post allogeneic SCT. Prior to lymphodepletion, 4 patients had >5% BM disease, 6 had disease between 5x10-2and 1x10-5, 4 were BM MRD negative having had recurrent isolated CNS disease. Median transduction efficiency was 31% (range 16.5 to 96.4%). 12/14 treated patients received the anticipated dose of 1x106CAR T cells/kg (2 received 0.9x106/kg). Considering all evaluable patients, (n=14 for CAR T cell persistence by qPCR, n=13 by flow) the geometric mean of Cmax was 128 912/µg DNA and of the area under the curve between D0 and D28 was 1,721,355 copies/ µg DNA (Table 1). At the point of maximal expansion, a median of 35% of circulating T cells were CAR+. Median half-life was 34 days (range 3-102). CAR T cells continued to be detectable by qPCR in 11 of 14 (79%) patients at last assessment and by flow cytometry up to 30 months post infusion in 8 of 13(61%). Median duration of CAR T persistence by flow was 261 days (range 7-917). 3 patients failed to have persistence of CAR T cells beyond 1 month. T cell mediated anti-CAR specific cytotoxic activity was detected in 2/2 evaluable patients. Updated persistence data will be presented at the meeting Cytokine release syndrome (CRS) occurred in 13 (93%, grade 1 n=9, grade 2 n=4). None developed ≥grade 3 CRS, had CRS-related ICU admission, or received Tocilizumab. CRS was associated with modest elevations of IL-6, IFN-γand IL-10. Grade 2 neurotoxicity was observed in 3 patients and resolved spontaneously. One patient had grade 4 leucoencephalopathy presumed due to chemotherapy as well as grade 5 sepsis. Ten patients (71%) had grade 3-4 cytopenia persisting beyond day 28 or recurring afterthis. 12/14 (86%) patients achieved molecular complete or continuing complete remission at a median of 30 days post infusion (range 30-90 days, Table 2). At a median follow-up of 20.3 months, 4/14 (29%) evaluable patients remain MRD negative. 5 relapsed with CD19-disease, 1 with CD19+ disease. The median duration of EFS (based on death or morphological relapse) has not been reached, 12 month EFS = 52%, OS = 70% (Figures 1, 2 and Table 3). Conclusion: We noted excellent CAR T cell expansion and persistence in a ALL cohort treated with the fast off-rate CAT-41BBz CAR despite their lower BM disease at treatment compared to other studies. The kinetics documented for all evaluable patients showed a 5-fold greater CAR T cell expansion and 2-fold longer half-life than responders in published series utilising tisagenlecleucel in a similar ALL cohort (Mueller et al., Blood 2017). Patients had a favourable toxicity profile with no severe (grade 3-4) CRS and equivalent disease outcomes to the ELIANA study despite having similarly advanced disease (Maude et al., NEJM 2018292). These data suggest long lived CAR T cell persistence supports stand-alone therapy for ALL with durable responses. Disclosures Ghorashian: Celgene: Honoraria; novartis: Honoraria; UCLB: Patents & Royalties: UCLB. Kramer:UCLB: Patents & Royalties. Ciocarlie:Servier: Other: Financial Support. Farzaneh:Autolus Ltd: Equity Ownership, Research Funding. Pule:Autolus: Employment, Equity Ownership, Patents & Royalties. Amrolia:UCLB: Patents & Royalties.

In search of the optimal platform for Post-Allogeneic SCT immunotherapy in relapsed multiple myeloma: a systematic review.

Allogeneic stem cell transplantation (allo-SCT) has the potential to induce sustained remissions in patients with multiple myeloma (MM). Currently, allo-SCT is primarily performed in high-risk MM patients, most often in the setting of early relapse after first-line therapy with autologous SCT. However, the implementation of allo-SCT for MM is jeopardized by high treatment-related mortality (TRM) rates as well as high relapse rates. In this systematic review, we aimed to identify a safe allo-SCT strategy that has optimal 1-year results regarding mortality, relapse and severe GvHD, creating opportunities for post-transplantation strategies to maintain remissions in the high-risk group of relapsed MM patients. Eleven studies were included. Median PFS ranged from 5.2 to 36.8 months and OS was 13.0 to 63.0 months. The relapse related mortality at 1 year varied between 0 and 50% and TRM between 8 and 40%. Lowest GvHD incidences were reported for conditioning regimens with T-cell depletion using ATG or graft CD34+ selection. Similar strategies could lay the foundation for a post-transplant immune platform, this should be further evaluated in prospective clinical trials.

A validated symptomatic BKV risk assessment score in patient post allogeneic stem cell transplant.

e18536 Background: BKV, Polyomavirus hominis 1, is a member of the family Polyomaviridae, is a non-enveloped virion; in the 1980s, it emerged as an important pathogen in SCT recipients. In the absence of sufficient T-cell immunity, BKV reactivation can progress, leading to prolonged hospital stays and increased mortality secondary to late hemorrhagic cystitis, ureteral stenosis, and nephropathy. In our recently completed retrospective study of 2477 SCT patients, 38.1% had developed renal impairment, and BKV viruria was present in 25%. In addition, BKV was found to be an independent predictor of chronic kidney disease and shorter survival. Using the large cohort (2477) patients studied earlier (2004-2012) we have derived a grading system to identify patients with risk of symptomatic BKV. We hypothesize that the current grading system will identify the patients at risk of symptomatic BKV at day 30 post allogeneic stem cell transplant. Methods: We performed a retrospective chart review of all patients who underwent allogeneic SCT from 2012-2016. The data was extracted from the secured database at MD Anderson cancer Center. Using the three variables that were significant predictor for symptomatic BKV derived from our initial study (conditioning regimen, HLA donor status, & underlying cancer diagnosis) we performed the analysis. Predicted cumulative incidence rate of BK infection at 30 days after transplant in 1308 patients were calculated in the presence of death as a competing risk using the “BASELINE” statement in PHREG procedure in SAS. Patients were classified into low, moderate and high risk according to the distribution of the predicted cumulative incidence of BK infection 30 days after transplant. Results: We have shown that the grading system derived from allogeneic SCT population predicted accurately the high, moderate & low risk population for developing symptomatic BKV. Conclusions: We have created and validated a grading system for symptomatic BKV in a large cohort of (1308 patients) to predict risk at day 30 post allogeneic SCT. Using this grading system we would hope to identify high risk patients for BKV and intervene early with novel therapies prior to complications associated.

Presence of GvHD and Fever of Unknown Origin Could be a Tubercular Infection Post Allogenic Hematopoietic Stem Cell Transplant

IntroductionPatients post allogeneic stem cell transplant (allo SCT) are expected to be at high risk of tuberculosis (TB) due to underlying immunosuppression. The incidence is predicted to be higher in countries endemic to TB, like ours. We conducted a retrospective study of post alloSCT patients for incidence of TB, its clinical presentation, factors associated with TB and its outcome.MethodsRecords of all patients who underwent allo SCT from 1st January 2012 till 31st December 2015 were reviewed for transplant related parameters and tubercular infection. The diagnosis of tuberculosis was considered if Mycobacterium tuberculosis (MTB) cultured from clinical samples or acid fast bacilli (AFB) demonstrated on histopathology/ smears. A possible TB was considered in presence of epitheloid cell granulomas without AFB. Patients already on anti-tubercular therapy (ATT) at the time of SCT or had any history of TB were excluded from study population.ResultsA total of 176 patients underwent allo SCT in study period with one patient having past history of TB lymphadenitis (excluded from further analysis). Indications for allo SCT were malignant diseases and non-malignant diseases in 82.3% (n=144) and 17.7% (n=31) patients respectively. Conditioning regimen was myeloablative in 57.7%(n=101) and reduced intensity in 20.6% (n=36) and non myeloablative in 21.7% (n=38). Total body irradiation (TBI) based myeloablative regimen was used in 24.5% (n=43) of patients. Anti-thymocyte globulin (ATG) was used in 14.8% of conditioning regimen. 19 patients underwent unrelated allotransplants. Overall 94 (53.7%) of patients experienced any graft vs host disease (GvHD) (acute 37% (n=65), chronic 29% (n=51) of all patients). Treatment related mortality in the study population was 26.3% (n=46) and overall survival was 50.3% (n=88) at 345 days of median follow up.Tuberculosis was detected in 5 patients (3 males, 2 females), none had a history of contact with TB, who underwent allo SCT (related =4, unrelated =1). Peripheral blood was used as graft source in all with myeloablative non TBI based conditioning regimen used in 2 patients while TBI based myeloablative conditioning regimen in 1 patient and reduced intensity conditioning in rest 2 patients. ATG was used in one patient for conditioning. Median ECOG performance score of all the 5 patients was 1 (0-1). Only one patient (diagnosis- acute lymphoblastic leukemia) received steroid based treatment as a part of pre transplant treatment during induction chemotherapy. Four patients developed pulmonary tubercular infection while one had lymph nodal involvement after allogeneic SCT, accounting to an incidence rate of tubercular infection of 2.8% at a median of 258 days post allo SCT. The presenting symptoms for TB were unexplained fever (n=4), weight loss (n=1), chest pain (n=2), cough with expectoration (n=1) which lasted for a median of 20 days till TB diagnosis was confirmed using lung biopsy (n=3), BAL culture (n =1) and lung FNA (n=1).All 5 patients had GvHD in one of its form and four patients who presented with pulmonary disease were receiving median number of 2 (1-3) immunosuppressive therapies at the time of TB diagnosis. Four patients with acute GvHD had received steroids as part of treatment. Presence of any GvHD and chronic GvHD was found to be significantly associated with Tubercular infection (p=0.042) and (p=0.002) respectively as per Fischer exact test. Acute GvHD, though present in 4/5 patients, had a trend towards Tubercular infection (p=0.058) (Table 1).All patients were started on 4 drug ATT with clinical/ radiological response in all. One patient developed transaminitis post 7 days of start of ATT for which dose modification was done and the other patient developed hyperbiluribinemia, which required temporary cessation of ATT. All the patients who developed TB are alive with a median overall survival of 1071 days post allo SCT.ConclusionThe incidence of TB is very high amongst allo SCT recipients, as compared to general population. Tuberculosis should be suspected in patients with unexplained fever post allogeneic SCT. Most of cases are pulmonary TB. Active GvHD and ongoing immunosuppression/ steroids are possible risk factors. Early diagnosis and treatment can salvage most of these patients and prevent long term morbidity and mortality. Hepatic toxicity following ATT is common and needs to be differentiated from hepatic GvHD. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Second Hematopoietic SCT for Thalassemia Major: Improved Clinical Outcomes with a Treosulfan Based Conditioning Regimen

Graft rejection (GR) following an allo-SCT occurs in 10-20% of patients with β thalassemia major (TM). The reported clinical outcome following second transplants have been associated with a high incidence of graft rejection, treatment related mortality, and graft versus host disease [Haematologica 2009; 94(9)]. There is limited data on the clinical profile and long term outcome of patients who have had a graft rejection. We undertook a retrospective analysis of patients who had a graft failure post allogeneic SCT for TM at our center.From October, 1991 to June 2016, 506 HLA matched related transplants for TM were done at our center. Of these 55 (11%) patients had a graft failure. An additional 7 patients with graft failure following an allo-SCT done at other centers were referred to us for a second transplant. Of the 62 patients with graft failure 32 (52.4%) were primary graft failures (PGF; 15 with aplasia and 17 with autologous recovery) while 30 (47.6%) were secondary graft failures (SGF; 5 with aplasia and 25 with autologous recovery). The median age of the patients who had graft failure was 8 years (range:1-19) and there were 38 (60.3%) males. On conventional risk stratification 40 (63.5%) were Class III. Eighteen (54.5%) cases with PGF and 16 (53.3%) with SGF did not receive a second transplant. From Oct 2009, at our center, a reduced toxicity myeloablative (MAC) treosulfan based conditioning regimen with a PBSC graft was offered to all high risk patients and for second transplants at the treating physician's discretion.Twenty nine (46%) of the patients with GR underwent a second allo-SCT. With the exception of one patient (first allo-SCT with an unrelated cord blood product) the donor for the second transplant was the same as the first transplant. Conditioning regimen for second SCT was busulfan based MAC in 7 (24%), treosulfan based MAC in 12 (41.3%) and the remaining received non-myeloablative conditioning regimens (fludarabine based, low dose TBI, OKT3, Cy-OKT3) often in view of pancytopenia and perceived inability to tolerate a MAC. All patients receiving a treosulfan based regimen had a PBSC graft. A BM graft was used in 7 (41%) of the remaining cases. None of the patients conditioned with a treosulfan based regimen had a graft rejection though one patient died of Grade IV acute GVHD. Of the remaining 17 patients 10 died following a second GR, 3 died of regimen related toxicity. Four are alive of which one has recurrent TM and the rest are well and transfusion independent at 55, 80 and 204 months from second transplant (all busulfan based MAC). The baseline characteristics and clinical outcomes of patients who received a treosulfan based MAC regimen versus the rest is summarized in table 1 and figure 1. On a univariate analysis a non-treosulfan based conditioning regimen and time from GR to second transplant of <1 year was significantly associated with an adverse impact. However, on a multivariate analysis only a non-treosulfan based regimen was associated with a significant adverse impact on EFS (HR=11.5; 1.13-116.4: P-value=0.039).In conclusion there has been a significant improvement in clinical outcomes in our experience with the use of a treosulfan based reduced toxicity MAC regimen for second allo-SCT for TM. It would be reasonable, where feasible, to defer the second transplant by a year following the first graft rejection. [Display omitted] [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Targeting Epigenetic Operations with HDAC Inhibitor and Hypomethylating Drugs in Combination Exhibit Synergy in Preclinical and Clinical Experiences in Drug Resistant T-Cell Lymphoma (TCL): A Translational Focus on Doublet Development

Introduction: The TCL are chemotherapy resistant neoplasms associated with a poor outcome. Recently, mutations in a number of important genes controlling epigenetic functions have been reported, including mutations in TET2, IDH1/2 and DNMT3. As a class, HDAC inhibitors have exhibited activity leading to regulatory approval only in TCL. Collectively, these observations suggest that combinations of hypomethylating agents (HoMe) and HDAC inhibitor (HDACi) should be explored as a potential new backbone in TCL. Methods : In vitro and in vivo models of TCL were used to evaluate the preclinical merits of combining HoMe and HDACi using standard cytotoxicity assays (CellTiter Glo and Anexin V staining) and well established murine NOG models. Gene expression array (GEP) using Affymatrix were performed to identify unique gene sets perturbed by the combination. Patterns of methylation and DNA fragmentation were determined using the LINE-1 Assay and APO alert DNA fragmentation assay . A Phase 1 study in patients with relapsed/refractory (R/R) lymphoma has enrolled all cohorts. Detailed PK and PD studies were performed. Results: In vitro , all 4 HDACi inhibitors (romidepsin [R]; vorinostat [V]; belinostat [B]; panobinostat [P]) exhibited marked class synergy with HoMe agents, including decitabine and azacytidine. Cell Titer Glo and Anexin V staining confirmed that the combination was potently synergistic as determined by the relative risk ratio (RRR). The synergy was more dependent on the concentration of the HDAC inhibitor, and was most pronounced with romidepsin versus other HDACi. In vivo , the combination of belinostat and decitabine produced a marked inhibition of TCL growth that was highly statistically significant compared to the single arm treated and untreated controls. GEP and methylation profiling revealed differentially expressed genes and modulated pathways for each of the single treatment conditions and combination. 944 unique genes were modulated by the combination supporting the idea of 9molecular synergism9. There was no methylation affect seen in the HDACi treated cells. Quantitation of DNA methylation and fragmentation suggested that concentrations of AZA from > 1 nM to Conclusions: A laboratory to bench translational initiative focused on identifying novel doublet combinations in TCL has confirmed that targeting epigenetic operations in TCL is effective in both the preclinical and clinical setting. Present efforts are directed towards accruing the Phase 2 component of the study in patients with PTCL, and completing all PK and PD studies from the Phase 1. We believe this combination could form the basis of a new backbone in TCL. Disclosures O9Connor: Seattle Genetics: Consultancy; Novartis: Consultancy, Honoraria; Bristol-Myers Squibb Company: Consultancy; Celgene: Consultancy, Research Funding; Takeda Millennium: Consultancy, Honoraria, Research Funding; Spectrum Pharmaceuticals: Consultancy, Honoraria, Research Funding; Mundipharma: Consultancy, Honoraria, Research Funding; Bayer: Consultancy, Honoraria; Acetylon Pharmaceuticals, INC: Consultancy. Off Label Use: Oral Azacitadine is not approved for lymphoid malignancies. Romidepsin is not approved for B-NHL and HL.. Deng: TG Therapeutics, Inc.: Honoraria, Research Funding; Seattle Genetics: Research Funding. Sawas: Seattle Genetics: Research Funding. Colbourn: Benitec Biopharma: Equity Ownership; PDL Biopharma: Equity Ownership; Merk: Equity Ownership; Eli Lilly: Equity Ownership; Celgene: Equity Ownership; Biogen Idecc: Equity Ownership; Novartis: Equity Ownership; Seattle Genetics: Equity Ownership. Neylon: Seattle Genetics: Speakers Bureau; Celgene: Speakers Bureau; Genentech: Speakers Bureau; Gilead: Speakers Bureau. Amengual: Acetylon Pharmaceuticals, INC: Consultancy, Research Funding.

Salvage therapy with everolimus reduces the severity of treatment-refractory chronic GVHD without impairing disease control: a dual center retrospective analysis.

Chronic GVHD (cGVHD) remains the most important cause of late non-relapse mortality post allogeneic hematopoietic SCT (HSCT). Although first-line treatment of cGVHD with steroids is well established, evidence for second-line treatment remains limited. Here, we report a dual center retrospective analysis of the off-label salvage treatment of steroid-refractory cGVHD with everolimus. Out of 80 patients with a median age of 50 (17-70) years, 14 (17%) suffered from mild, 39 (49%) from moderate and 27 (34%) from severe cGVHD. At the final analysis, median follow-up after introduction of everolimus was 724 (14-2205) days. Thirty-four patients (43%) required the addition of further immunosuppression during everolimus-based therapy. Global NIH Severity Score improved in 34 patients (43%), remained stable in 37 patients (46%) and worsened in 9 patients (11%). The total sum of Global NIH Severity Scores in all patients assessable was significantly reduced after treatment with everolimus (P<0.0001). Most frequent grade 3/4 toxicities included infections (n=30) and thrombocytopenia (n=15). There was a single case of relapse. Everolimus-based salvage treatment of refractory cGVHD results in significant improvement of the NIH Severity Score without impairing control of the malignant disease. Finally, these preliminary results demand further verification in prospective trials.

Differences in health behaviour between recipients of allogeneic haematopoietic SCT and the general population: a matched control study.

Little is known of health-relevant behaviour among long-term survivors of haematological disorders treated with haematopoietic SCT. This comparative cross-sectional multicentre study aimed (1) to explore the prevalence of selected behaviours in this group and (2) to compare them with those of the general population. Self-reported data of 376 survivors (mean age: 50.4 (s.d. = 12.8); median 7 years postallogeneic SCT (interquartile range (IQR) = 8.9; range 1-33) were compared with controls derived from the Swiss Health Survey 2007 by propensity score matching. Survivors were more physically inactive (26.8% vs 12.5%; P ⩽ 0.001) and consumed fewer portions of vegetables (⩾ 3 pieces: 10% vs 21.6%; P < 0.001), fruits (⩾ 3 pieces: 6.5% vs 10.6%; P < 0.001) and fish (31.2% vs 60.9% weekly fish dish; P < 0.001). More survivors consumed dairy products daily (92.5% vs 62.9%; P < 0.001), used sun protection regularly (94.5% vs 85.3%, P < 0.001) and had received influenza vaccinations in the past year (58.4% vs 21.5%; P < 0.001); fewer survivors smoked (13.4% vs 35.4%; P < 0.001). Survivors' weekly alcohol consumption was lower (median 1.5 servings (IQR 4) vs median 4.5 (IQR 10.3); P < 0.001). Of those taking immunosuppressants, 65.7% were non-adherent. Similar to the general population, survivors experience problems executing several health-enhancing behaviours, warranting corrective interventions.

Severe lung injury and lung biopsy in children post‐hematopoietic stem cell transplantation: The differences between allogeneic and autologous transplantation

To review outcome of children post-allogeneic (allo) and autologous (auto) SCT with severe lung injury who had lung biopsy and to determine whether the diagnoses provided by lung biopsy had an impact on outcome. Retrospective study was carried out from January 2000 to June 2010. Nine hundred and eighteen children (0-18yr) received SCT (allo 476, auto 442), and 59 biopsies were performed in 48 patients. Most common result of lung biopsy was non-infectious inflammation and recurrent disease in allo- and autorecipients, respectively. In a multivariate analysis, survival of allorecipients who had management change was inferior (p=0.002; HR: 3.12). These patients were extremely sick, and management change was the last attempt to stabilize their respiratory status. There was a trend toward superior survival for children who had biopsy after 100days following SCT (p=0.09; HR: 0.55) and a trend toward inferior survival for those with proven infections within twowk of biopsy (p=0.07; HR: 2.14). Only 31% of allorecipients and 25% of autorecipients survived. There were no biopsy-related complications. Lung biopsy itself appears to be well tolerated, although requiring a biopsy seems to carry a poor prognosis; this seems to be due to different causes, auto (relapse), allo (non-infectious inflammation).

Viral-specific adoptive immunotherapy after allo-SCT: the role of multimer-based selection strategies

Recipients of hematopoietic SCT undergo a period of profound immunosuppression due to the chemotherapy and/or radiotherapy used for the conditioning and to the graft versus host reaction. SCT patients are highly susceptible to the development of viral infections such as CMV or EBV. The achievement of a competent immunological response, such as viral-specific T cells, is associated with a lower incidence of viral infections. Methods for direct identification of antigen-specific T cells have been based on the functional characteristics of these T cells. Techniques such as proliferation and ELISPOT assays, intracellular cytokine staining and IFN-γ capture have been used to quantitate and obtain viral-specific T cells. Multimers are composed of several MHC molecules loaded with immunodominant peptides joined to a fluorescent molecule, which signal can be quantified by a flow cytometer. Multimer technology together with recent advances in flow cytometry, have facilitated the monitoring and selection of antigen-specific T cells without the need for in vitro cultures and manipulation. This has resulted in a better characterization of the function and phenotype of the different subpopulations of T cells involved in the immune recovery post allogeneic SCT. It is becoming a distinct possibility to isolate individual antigen-specific T cells, without long-term culture techniques, and potentially use them as adoptive immunotherapy in the SCT setting.

Long-Term Disease-Free Survival of Patients with AML Relapse After T-Cell-Depleted Allogeneic Stem Cell Transplantation by Re-Induction Therapy and Subsequent Interferon-Boosted Donor Lymphocyte Infusion

Abstract 3064Relapse of AML after allogeneic stem cell transplantation (alloSCT) has a very poor prognosis. Salvage re-induction chemotherapy leads to clinical remissions in a substantial number of patients, but these remissions tend to be of short duration. In contrast, donor lymphocyte infusions (DLI) have the potential to effect long-lasting remissions, but the interval of several weeks to months that is required to develop a DLI-induced anti-leukemia response may prevent efficient control of a highly proliferative leukemia. In addition, a high tumor burden may suppress the immune response. In contrast, the combination of efficient cytoreduction by chemotherapy with DLI administered in rapid succession under circumstances favoring the development of an early and profound immune response might have the potential to eradicate otherwise resistant leukemia cells. We therefore adopted an institutional therapeutic strategy for relapsed myeloid leukemia post-allogeneic SCT based on administration of DLI at the anticipated end of the neutropenic phase after salvage re-induction chemotherapy. At this time point, the high prevalence of a pro-inflammatory milieu should favor the induction of the immune response, and an expected state of lymphopenia should promote the expansion of infused T cells by homeostatic proliferation. If 3 weeks after DLI no graft versus host disease (GvHD) was observed, the potential anti-leukemia immune response was further amplified by treatment with interferon- α (IFN- α) until GvHD occurred.Between January 2000 and December 2009 44 patients with relapsed myeloid malignancy after alloSCT were treated at our hospital. Pre-transplant diagnoses were AML n=40, CMML n=1 and MDS n=3. Median time from SCT to relapse was 187 days. Median follow-up after relapse was 3.1 years. 5 patients had a smouldering relapse (<10% bone marrow blasts) and 39 patients had an overt relapse.Of 39 patients with overt relapse, 7 patients (18%) did not receive re-induction therapy due to poor performance status (n=5) or patient choice (n=2). 32 patients received remission-induction therapy consisting of gemtuzumab ozogamycin (n=9), cytosine arabinoside-containing chemotherapy (n=17), or both (n=6). Following this treatment, 7 of 32 patients had rapidly progressive disease during induction therapy (n=6) or died due to toxicity (n=1) and did not receive DLI.The remaining 25 patients received DLI at a dose of 5.0×10 ^6 CD3+ cells/kg for related and 2.5×10 ^6 CD3+ cells/kg for unrelated donors 3 weeks after the start of remission-induction therapy. In 16 of these patients DLI was boosted with IFN- α 3.0×10 ^6 IE once daily. This strategy resulted in acute GvHD in 17 of 25 patients (n=8 grade 1–2, n=9 grade 3–4). At 6 weeks after DLI, 16 patients had reached CR, 5 patients had failed to reach CR (2 with GvHD) and 4 suffered treatment-related mortality (3 with GvHD). Of the 16 patients in CR, 4 had no signs of GvHD and developed a second relapse during the follow-up period. Only 3 of 12 patients in CR with signs of acute GvHD at 6 weeks after DLI developed a second relapse. In total, 9 of 17 patients (53%) with acute GvHD after DLI had long term survival versus none without acute GvHD. During follow-up, 8 patients developed chronic GvHD (n=4 limited, n=4 extensive).Finally, 5 patients with an early detected smouldering relapse received DLI, which was boosted with IFN- α in 2 patients, without salvage re-induction therapy. All 5 patients developed GvHD (n=2 grade 1–2, n=3 grade 3–4) and 3 patients achieved a CR of whom 1 patient died from GvHD.Our results indicate that treatment of relapsed AML after alloSCT with salvage re-induction therapy followed by DLI at the end of the neutropenic phase during minimal residual disease, with additional boosting of the immune response with IFN- α, can result in long-term disease-free survival. Disclosures:Off Label Use: Interferon: DLI-boosting.

Recombinant human soluble thrombomodulin for the treatment of hepatic sinusoidal obstructive syndrome post allogeneic hematopoietic SCT

Recombinant human soluble thrombomodulin for the treatment of hepatic sinusoidal obstructive syndrome post allogeneic hematopoietic SCT

Assessment of the Impact of Human Herpesvirus 6 (HHV-6) Replication On Active Citomegalovirus (CMV) Infection After Allogeneic Stem Cell Transplantation (Allo-SCT).

Abstract 2225Poster Board II-202 Introduction:In spite of significant advances in the diagnosis and treatment of CMV infection post-allogeneic SCT it continues to be associated with significant morbidity and mortality. The human herpesvirus 6 (HHV-6) reactivation has been associated with different complications, including delayed neutrophil and platelet engraftment, interstitial pneumonia, skin rash, severe graft versus host disease (GvHD), and central nervous system disorders. HHV-6 is considered an immunomodulatory and immunosuppressive agent, and thereby, it may the increase the risk of active CMV infection and disease in the transplantation setting Aims:To determine if the HHV-6 facilitates the CMV reactivation or influences the kinetic of CMV replication and its specific immune replication during the first 100 days after allogeneic hematopoietic SCT. Patients and Methods:Between Nov-05 and Feb-08, sixty-eight patients were monitored for plasma HHV-6 and CMV DNAemia by PCR once/week until day +100 after Allo-SCT. Characteristics of patients: median age: 45 yo (18-70); sex (M/F): 37/31. Diagnosis: AML/MDS(n=27), ALL(n=10), lymphoproliferative syndrome(n=17), MM (n=4), CML (n=3), Others(n=7). CMV seropositive (R/D)(%): 88/60. Cell source: PBSC: 53(78%);CB:13(19%);BM:2(3%). Conditioning treatment: RIC: 40 pts, myeloablative: 28 pts. pp65 antigenemia in PMN leucocytes (Light Diagnostics, Chemicon Intern). Positive if ≥1cel+/200.000. Plasma DNA-CMV: PCR-RT Abbott or Amplicor CMV Monitor Roche. Plasma DNA-HHV6: Q-PCR Alert Amplimix, Nanogen Adv. Diag., detection limit: 10 copies/mL plasma. Enumeration of pp65 and IE-1 IFNγ CD4+ and CD8+ T cells was performed by intracellular cytokine staining. (BD Fastimmune, BD-Beckton Dickinson). Pre-emptive antiviral treatment was initiated if pp65 Ag +. Results:HHV-6 DNAemia occurred in 27 out of 68 pts (40%), after a median time of 20 days post-SCT with a peak value of 345 copies (26-13,661) reached on the fourth week in the 52% of patients, with a median duration of 10 days (3-35). Twenty-three out of the 27 patients were HHV-6 seropositive before transplantation. Clearance of most episodes (21 out of 27) occurred in the absence of (val)ganciclovir treatment, which was initiated in the course of the remaining 6 episodes because of the development of antigenemia-positive active CMV infection. HHV-6 DNAemia was significantly associated with subsequent CMV DNAemia in univariate (P=.01), but not in multivariate analysis (P=.056) that included transplant from URD, HLA non identical donor, myeloablative conditioning, CBT, use of prednisone as GVHD prophylaxis and plasma HHV-6 DNA detection. HHV-6 DNAemia was not predictive of development of CMV DNAemia. Timing and kinetics of active CMV infection were comparable in patients either with or without a preceding episode of HHV-6 DNAemia. Occurrence of HHV-6 DNAemia had no impact on the level of CMV-specific T-cell immunity reconstitution early after transplant. The receipt of a graft from an unrelated donor was the principal variable associated with HHV-6 and CMV coactivation. Conclusion:The results of this study suggest that a basal immunosuppressive state, specially the receipt of a graft from an unrelated donor, leads to HHV-6 and CMV coactivation, and argue against an effect of active HHV-6 infection on CMV replication either by a direct interaction or indirectly by suppressing CMV-specific T-cell responses. Disclosures:No relevant conflicts of interest to declare.

Long-term outcome after allo-SCT: close follow-up on a large cohort treated with myeloablative regimens

We analyzed the late outcomes of 429 long-term survivors post allogeneic hematopoietic SCT (allo-HSCT) who received transplant in our center between 1981 and 2002, and were free of their primary disease for > or =2 years after allo-HSCT. Late recurrent primary malignancy was found in 58 (13.5%) patients and was the primary cause of late death. A total of 37 (8.6%) patients died of non-relapse causes at a median of 5.5 years (range, 2-15.6 years) post allo-HSCT. The major non-relapse causes of death were chronic GVHD (cGVHD), secondary malignancy and infection. The probabilities of OS and EFS were 85% (95% cumulative incidence (CI) (81-89%)) and 79% (95% CI (74-83%)) at 10 years, respectively. Long-term allo-HSCT survivors were evaluated for late complications (median follow-up, 8.6 years (range, 2.3-22.8 years)). cGVHD was diagnosed in 196 (53.1%) survivors. The endocrine and metabolic complications were hypogonadism in 134 (36.3%) patients, osteopenia/osteoporosis in 90 (24.4%), dyslipidemia in 33 (8.9%), hypothyroidism in 28 (7.6%) and diabetes in 28 (7.6%). Hypertension was diagnosed in 79 (21.4%), renal impairment in 70 (19.0%), depression in 40 (10.8%) and sexual dysfunction in 33 (8.9%) survivors. We conclude that in patients who receive allo-HSCT as treatment for hematological malignancy and who are free of their original disease 2 years post transplant, mortality is low and the probability of durable remission is high. Lifelong surveillance is recommended.

Delay in B-lymphocyte recovery and function following rituximab for EBV-associated lymphoproliferative disease early post-allogeneic hematopoietic SCT

Treatment with rituximab is highly effective for EBV-associated post transplant lymphoproliferative disease. However, little is known about its immunological sequelae in pediatric allogeneic hematopoietic SCT (HSCT). Time to normal CD19+ B-lymphocyte values in blood and intravenous immunoglobulin (IVIG) substitution needed to maintain an IgG>400 mg per 100 ml in six consecutive pediatric allogeneic HSCT patients treated with rituximab for symptomatic EBV reactivation were compared with a matched cohort of non-rituximab-treated patients. Follow-up of the six patients ranged from 149 to 1546 days; all but one survived. The mean (+/-s.d.) time to recovery of CD19+ B-lymphocytes was 353+/-142 days as compared with 139+/-42 in the controls (P<0.01). Similarly, substitution of IVIG as a measure of functional B-cell recovery was extended from a mean of 122+/-45 to a mean of 647+/-320 days, and the cumulative dose of IVIG increased from a mean of 1.86+/-0.51 to 4.4+/-0.97 g/kg, respectively (P<0.05). One patient had functional B-lymphocyte deficiency for >3 years and ultimately required two stem cell boosts. Rituximab is a live-saving treatment for pediatric HSCT patients but may lead to prolonged and even persistent B-cell deficiency.

In vivo expansion of LMP 1- and 2-specific T-cells in a patient who received donor-derived EBV-specific T-cells after allogeneic stem cell transplantation

Immunotherapy approaches with antigen-specific cytotoxic T lymphocytes (CTLs) have provided safe and effective prophylaxis and treatment of Epstein-Barr virus (EBV)-associated lymphomas arising after bone marrow transplantation. EBV is also associated with other malignancies including ∼40% of cases of Hodgkin's disease, making this tumor another potential target for EBV-targeted immunotherapy. This study describes a patient with multiple relapsed EBV positive Hodgkin's Disease who received both autologous and allogeneic EBV CTL lines. After multiple chemotherapeutic and radiotherapy regimens including two autologous stem cell transplants, he received two doses of gene-marked autologous EBV-specific CTL which resulted in disease stabilization for 6 months. The gene-marked EBV-CTL persisted for 12 months in the peripheral blood after which he proceeded to unrelated donor stem cell transplant followed by immunotherapy with donor-derived EBV-specific CTL. Despite low levels of donor chimerism, the patient remains in complete remission 5 years post-allogeneic SCT. Comparison of the autologous and the donor-derived CTL lines showed that the donor line had specificity for two tumor-associated EBV antigens, latent membrane protein (LMP)1 and 2 compared to the autologous line, which only had specificity for LMP2 epitopes. Following infusion of the donor-derived CTL, functional analyses showed that T-cells reactive with both LMP1 and LMP2 epitopes expanded in the peripheral blood, suggesting that strategies to increase their frequency may result in a broader cytotoxic response against EBV+ Hodgkin tumors.