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BACKGROUNDLeukemia recurrence is a major cause of treatment failure after autologous stem cell transplantation for acute myeloid leukemia (AML). It usually occurs within the first 2 years after transplantation. The goal of the current retrospective study was to assess the follow-up of and characterize risk factors for outcome among patients who survived free of disease recurrence after this period. METHODSThe analysis included 3567 adults (median age, 45 years) with AML who underwent autografting during the first (86% of patients) or second (14% of patients) complete remission between 1990 and 2008. The stem cell source was the bone marrow in 32% of patients or the peripheral blood in 68% of patients. The median follow-up was 6.9 years. RESULTSAt 5 years and 10 years after transplantation, the probability of leukemia-free survival was 86% and 76%, respectively; the recurrence incidence was 11% and 16%, respectively; and the nonrecurrence mortality rate was 3% and 8%, respectively. The observed survival was decreased compared with the expected survival of the general European population. In a multivariate analysis, decreased probability of leukemia-free survival was demonstrated for patients who underwent peripheral blood autologous stem cell transplantation; had French-American-British subtypes M0, M6, or M7; and were of an older age. The same factors were found to be associated with an increased risk of disease recurrence. Nonrecurrence mortality was found to be affected by older age. CONCLUSIONSThe results of the current analysis indicate that late recurrences remain a major concern after autologous stem cell transplantation among patients with AML, indicating the need for close monitoring of minimal residual disease and additional leukemic control measures after transplantation. Cancer 2016;122:1880-7. (c) 2016 American Cancer Society. Late disease recurrence remains a major concern for long-term survivors of autologous stem cell transplantation for acute myeloid leukemia. This indicates the need for close monitoring of minimal residual disease and additional leukemic control measures after transplantation.

Item does not contain fulltext Steroid-refractory acute graft-versus-host disease (aGVHD) remains an important cause of morbidity and mortality after allogeneic stem cell transplantation (SCT). A protocol on the management of aGVHD was introduced in our center that incorporated a prospective study on combination therapy with inolimomab (anti-IL-2Ralpha) and etanercept (anti-tumor necrosis factor-alpha) for steroid-refractory aGVHD. We evaluated the efficacy and safety in 21 consecutively treated patients. The patients had developed refractory aGVHD after SCT (n = 16) or donor lymphocyte infusion (n = 5), and aGVHD was classified as severe in all patients, mostly due to gastrointestinal involvement stages 2 to 4. No drug-related side effects were observed apart from the infections expected to occur in these severely immunocompromised patients. Overall response at day 28 of second-line therapy was 48% (10/21), with 6 and 4 patients achieving a complete and partial response, respectively. Eventually, 19 patients died (90%), with early mortality (

Item does not contain fulltext We describe the use and outcomes of allogeneic hematopoietic stem cell transplantation (allo-HSCT) for multiple myeloma (MM) in Europe between January 1990 and December 2012. We identified 7333 patients, median age at allo-HSCT was 51 years (range: 18-78), of whom 4539 (62%) were males. We distinguished three groups: (1) allo-HSCT upfront (n=1924), (2) tandem auto-allo-HSCT (n=2004) and (3) allo-HSCT as a second line treatment and beyond (n=3405). Overall, there is a steady increase in numbers of allo-HSCT over the years. Upfront allo-HSCT use increased up to year 2000, followed by a decrease thereafter and represented 12% of allo-HSCTs performed in 2012. Tandem auto-allo-HSCT peaked around year 2004 and contributed to 19% of allo-HSCTs in 2012. Allo-HSCT as salvage after one or two or three autografts was steadily increasing over the last years and represented 69% of allo-HSCTs in 2012. Remarkable heterogeneity in using allo-HSCT was observed among the different European countries. The 5-year survival probabilities from time of allo-HSCT for the three groups after year 2004 were 42%, 54% and 32%, respectively. These results show that the use of allo-HSCT is increasing in Europe, especially as second line treatment and beyond. There is an unmet need for well-designed prospective studies investigating allo-HSCT as salvage therapy for MM.

Item does not contain fulltext Dendritic cell (DC)-based vaccination boosting antigen-specific immunity is being explored for the treatment of cancer and chronic viral infections. Although DC-based immunotherapy can induce immunological responses, its clinical benefit has been limited, indicating that further improvement of DC vaccine potency is essential. In this study, we explored the generation of a clinical-grade applicable DC vaccine with improved immunogenic potential by combining PD-1 ligand siRNA and target antigen mRNA delivery. We demonstrated that PD-L1 and PD-L2 siRNA delivery using DLin-KC2-DMA-containing lipid nanoparticles (LNP) mediated efficient and specific knockdown of PD-L expression on human monocyte-derived DC. The established siRNA-LNP transfection method did not affect DC phenotype or migratory capacity and resulted in acceptable DC viability. Furthermore, we showed that siRNA-LNP transfection can be successfully combined with both target antigen peptide loading and mRNA electroporation. Finally, we demonstrated that these PD-L-silenced DC loaded with antigen mRNA superiorly boost ex vivo antigen-specific CD8(+) T cell responses from transplanted cancer patients. Together, these findings indicate that our PD-L siRNA-LNP-modified DC are attractive cells for clinical-grade production and in vivo application to induce and boost immune responses not only in transplanted cancer patients, but likely also in other settings. 01 februari 2013

Item does not contain fulltext Graft-versus-host disease (GVHD) and fungal infections are frequent complications after allogeneic hematopoietic stem cell transplantation (HSCT). Single nucleotide polymorphisms (SNPs) in genes of the immune system can influence the inflammatory cascade and T cell-driven alloimmune reactions after HSCT, and thus increasing the incidence of GVHD and infectious complications. Here, we investigated the effect of SNPs in IL-23R and CCR6 on posttransplantation outcome in 161 recipients of partially T cell-depleted HSCT. Remarkably, IL-23R SNPs were not associated with clinical outcome, but we found that disparities in the CCR6 tagSNP rs2301436 and SNP rs3093023 are independently associated with the occurrence of chronic GVHD (cGVHD) and invasive fungal disease. In multivariate analysis, patients receiving a transplant from a homozygous rs2301436 G allele donor showed less cGVHD (odds ratio [OR]: 0.16; P = .002), as was the case for a homozygous donor rs3093023 G allele (OR: 0.24; P = .005). In parallel, the GG genotype at rs2301436 in donors was associated with a higher incidence of invasive fungal disease at day 100 after HSCT (OR: 3.59; P = .008). This study shows that CCR6 SNPs can be used to predict clinical outcome, and that polymorphisms in the CCR6 gene may influence T cell-mediated immune reactions after HSCT.

Contains fulltext : 97194.pdf (Publisher’s version ) (Closed access) Tumor relapses remain a serious problem after allogeneic stem cell transplantation (alloSCT), despite the long-term persistence of minor histocompatibility antigen (MiHA)-specific memory CD8(+) T cells specific for the tumor. We hypothesized that these memory T cells may lose their function over time in transplanted patients. Here, we offer functional and mechanistic support for this hypothesis, based on immune inhibition by programmed death-1 (PD-1) expressed on MiHA-specific CD8(+) T cells and the associated role of the PD-1 ligand PD-L1 on myeloid leukemia cells, especially under inflammatory conditions. PD-L1 was highly upregulated on immature human leukemic progenitor cells, whereas costimulatory molecules such as CD80 and CD86 were not expressed. Thus, immature leukemic progenitor cells seemed to evade the immune system by inhibiting T-cell function via the PD-1/PD-L1 pathway. Blocking PD-1 signaling using human antibodies led to elevated proliferation and IFN-gamma production of MiHA-specific T cells cocultured with PD-L1-expressing leukemia cells. Moreover, patients with relapsed leukemia after initial MiHA-specific T-cell responses displayed high PD-L1 expression on CD34(+) leukemia cells and increased PD-1 levels on MiHA-specific CD8(+) T cells. Importantly, blocking PD-1/PD-L1 interactions augment proliferation of MiHA-specific CD8(+) memory T cells from relapsed patients. Taken together, our findings indicate that the PD-1/PD-L pathway can be hijacked as an immune escape mechanism in hematological malignancies. Furthermore, they suggest that blocking the PD-1 immune checkpoint offers an appealing immunotherapeutic strategy following alloSCT in patients with recurrent or relapsed disease.

Rationale: FLT3 in mutated in 30% of AML and is associated with poor prognosis. Allogeneic hematopoietic cell transplantation (allo-HCT) in first complete remission (CR) is recommended in FLT-3 ITD AML. However frequent and early post-transplant relapse leads to poor outcome. Several small studies suggested the efficacy of sorafenib as prophylactic or preemptive therapy or as treatment for relapse post allo-HCT. The purpose of this study was to assess the impact of sorafenib on outcomes of FLT3 mutated AML post allo-HCT. Patients and Methods: We identified 462 adult patients (51% males) with FLT3 mutated AML (FLT3 ITD-95%) allografted at EBMT participating centers between 2010 and 2015. Median age was 50 years (range 18-75). Patients mostly belonged to the intermediate cytogenetic risk group (82%) and 55% of those with available data had NPM1 mutation. Disease status at allo-HCT was CR1 (72%), CR2 (10%) or active disease (18%). Donors were matched related (MSD, 40%), matched unrelated (49%) or haploidentical (11%), respectively. Preparative regimens were myeloablative (MAC) in 53% and reduced intensity conditioning (RIC) in 47% of allo-HCT. Sixty two patients received post-transplant sorafenib either as prophylactic (n=19) or preemptive therapy (n=9) or as treatment for relapse (n=34). Median time from allo-HCT to initiation of maintenance sorafenib was 55 days (1-173). Median follow-up for alive patients was 39 months (range 1-87). Results: The 2-year leukemia free survival (LFS), overall survival (OS) and GVHD relapse free survival (GRFS) was 51%, 59% and 38%, respectively. In multivariate Cox analysis, sorafenib maintenance (either prophylactic or preemptive) significantly improved OS (HR=0.36; p=0.03). OS was also positively affected by NPM1 mutation, allo-HCT in CR1, and in vivo T cell depletion, but negatively affected by the need for >1 induction. Similarly, sorafenib maintenance significantly improved GRFS (HR=0.44; p=0.02). GRFS was also positively affected by NPM1 mutation (HR=0.66; p=0.002), haploidentical donors compared to MSD (HR=0.61; p=0.04), and in vivo T cell depletion (HR=0.55; p=0.00001), but negatively affected by the need for >1 induction (HR=1.5; p=0.005) or active disease at transplant (HR=2.5; p than 50 years), disease status at allo-HCT (CR1 Vs CR2 Vs active disease), conditioning (RIC Vs MAC), and time from allo-HCT to relapse (+/- 1 month). One year and 2 year OS were 41% and 30% for patients treated with sorafenib versus 14% and 14% for controls (P=0.0015). Conclusion: For AML patients with FLT3 mutation, post-transplant sorafenib whether given as prophylactic or preemptive therapy or as treatment for relapse, significantly improves OS and may be considered as standard of care in that setting. Disclosures Mohty: MaaT Pharma: Consultancy, Membership on an entity's Board of Directors or advisory committees; Servier: Consultancy; Jazz Pharmaceuticals: Honoraria, Research Funding, Speakers Bureau; Bristol Myers: Consultancy, Research Funding; Janssen: Honoraria, Research Funding, Speakers Bureau; Molmed: Consultancy; Sanofi: Consultancy, Honoraria, Research Funding, Speakers Bureau; Celgene: Consultancy, Honoraria; Amgen: Consultancy, Honoraria; Takeda: Honoraria, Speakers Bureau.

Contains fulltext : 89843.pdf (Publisher’s version ) (Closed access) Allogeneic stem cell transplantation (SCT) in multiple myeloma (MM) may induce a curative graft-versus-myeloma (GVM) effect. Major drawback in unmanipulated reduced-intensity conditioning (RIC) SCT is the risk of severe and longstanding graft-versus-host-disease (GVHD). This study demonstrates that transplantation with a partial T cell-depleted graft creates a platform for boosting GVM immunity by preemptive donor lymphocyte infusion (DLI) and recipient dendritic cell (DC) vaccination, with limited GVHD. All 20MM patients engrafted successfully. Chimerism analysis in 19 patients evaluable at 3 months revealed that 7 patients were complete donor, whereas 12 patients were mixed chimeric. Grade II acute GVHD (aGVHD) occurred in 7 patients (35%) and only 4 patients (21%) developed chronic GVHD (cGVHD). Fourteen patients received posttransplantation immunotherapy, 8 preemptive DLI, 5 patients both DLI and DC vaccination, and 1 patient DC vaccination only. DC vaccination was associated with limited toxicity, and none of these patients developed GVHD. Importantly, overall treatment-related mortality (TRM) at 1 year was low (10%). Moreover, the overall survival (OS) is 84% with median follow-up of 27 months, and none of the patients died from progressive disease. These findings illustrate that this novel approach is associated with limited GVHD and mortality, thus creating an ideal platform for adjuvant immunotherapy. 01 maart 2010

Adoptive T cell therapy has shown clinical potential for patients with cancer, though effective treatment is dependent on longevity and potency of the exploited tumor-reactive T cells. Previously, we showed that ex vivo inhibition of AKT using the research compound Akt-inhibitor VIII retained differentiation and improved functionality of minor histocompatibility antigen (MiHA)-specific CD8+ T cells. Here, we compared a panel of clinically applicable AKT-inhibitors with an allosteric or adenosine triphosphate-competitive mode of action. We analyzed phenotype, functionality, metabolism and transcriptome of AKT-inhibited CD8+ T cells using different T cell activation models. Most inhibitors facilitated T cell expansion while preserving an early memory phenotype, reflected by maintenance of CD62L, CCR7 and CXCR4 expression. Moreover, transcriptome profiling revealed that AKT-inhibited CD8+ T cells clustered closely to naturally occurring stem cell-memory CD8+ T cells, while control T cells resembled effector-memory T cells. Interestingly, AKT-inhibited CD8+ T cells showed enrichment of hypoxia-associated genes, which was consistent with enhanced glycolytic function. Notably, AKT-inhibition during MiHA-specific CD8+ T cell priming uncoupled preservation of early memory differentiation from ex vivo expansion. Furthermore, AKT-inhibited MiHA-specific CD8+ T cells showed increased polyfunctionality with co-secretion of IFN-γ and IL-2 upon antigen recall. Together, these data demonstrate that AKT-inhibitors with different modality of action promote the ex vivo generation of stem cell memory-like CD8+ T cells with a unique metabolic profile and retained polyfunctionality. Akt-inhibitor VIII and GDC-0068 outperformed other inhibitors, and are therefore promising candidates for ex vivo generation of superior tumor-reactive T cells for adoptive immunotherapy in cancer patients.

Contains fulltext : 89048.pdf (Publisher’s version ) (Closed access) Primary myelofibrosis is a clonal haematopoietic stem cell disease, characterised by marrow stromal fibrosis, extramedullary haematopoiesis, splenomegaly, hepatomegaly and progressive cytopenia. Therapeutic options once cytopenia has developed are limited to supportive care, such as erythrocyte transfusions and growth factors. The aetiology has become more clear, especially since JAK-2 mutations were found, resulting in increased production of cytokines. The immune-modulating drug thalidomide and its derivative lenalidomide have shown to be effective in reducing cytopenia, most probably by inhibiting the cytokine responses. In some patients the bone marrow fibrosis disappears. We describe the experience with these drugs in a cohort of 14 patients for thalidomide and seven for lenalidomide (in six patients lenalidomide was given after thalidomide and one patient received lenalidomide upfront). Thalidomide gave clinical improvement in 6/14 patients, but its use was limited mainly due to toxicity, especially the development of neuropathy. The drug could be given for a median period of 15.5 months in responding patients. Lenalidomide was effective in 4/7 of the patients, in some patients with no response on thalidomide. Due to the more favourable toxicity profile, the median duration of therapy was 19 months, with 3/4 patients on therapy longer than 19 months. These data are discussed in view of the clinical studies published. We conclude that lenalidomide is preferred in myelofibrosis, given a higher response rate and more favourable toxicity profile. If no response the addition of prednisone can be considered. In some patients it can normalise haemoglobin and make them transfusion independent. 01 augustus 2010

Contains fulltext : 69575.pdf (Publisher’s version ) (Closed access) BACKGROUND: Alloreactive NK cells play a role in tumor eradication after allogeneic HLA mismatched stem cell transplantation (SCT). The effect of NK alloreactivity in HLA identical SCT is still under debate and in particular in transplantation for chronic myeloid leukemia (CML) the data are very limited and with conflicting outcome. The aim of our study was to evaluate the effect of KIR genes and KIR ligands on leukemia free survival (LFS) and relapse rate in a well-defined, homogeneous group of CML patients phase upon HLA identical sibling SCT. METHODOLOGY: We retrospectively analyzed the effect of KIRs and KIR ligands (C1 and C2) on LFS and relapse in 70 CML patients in 1st chronic phase, who had received an HLA identical sibling graft. For KIR typing we used a single PCR based KIR typing protocol that also included primers allowing for the identification of the KIR binding site on HLA-Cw (AA 77 and 80). PRINCIPAL FINDINGS: The data show clear differences in transplant outcome between patients having both ligands (C1 and C2) as compared to patients having only one ligand (C1 or C2). In the latter group, the stimulatory KIR2DS5 gene was associated with improved leukemia free survival (p=0.007; hazard ratio 4.3; 95% confidence interval 1.3-6.7) and lower relapse rates (p=0.028; HR 4.3, 95% CI 1.1-9.1). In contrast, in patients carrying both ligands, KIR2DS5 was associated with reduced LFS (p=0.0056; HR 0.3; 95% CI 0.1-0.7) and higher relapse rate (p=0.02; HR 0.35, 95% CI 0.1-0.8). CONCLUSIONS: Our data indicate a role for an NK mediated anti-CML response after HLA identical sibling SCT that is influenced by KIR ligands and, more importantly, by stimulatory KIRs present in the donor.

Contains fulltext : 52018.pdf (Publisher’s version ) (Closed access) BACKGROUND: In trauma, as interventions are carried out to stop bleeding, ongoing resuscitation with blood products is of vital importance. As transfusion policy in exsanguinating patients cannot be based on laboratory tests, transfusion of blood products is performed empirically or 'blindly'. The aim of this study was to delineate 'blind' transfusion practice in the hectic clinical situation of exsanguination. METHODS: Seventeen trauma patients were selected who died due to uncontrolled bleeding despite haemostatic interventions within 24h after admission and who received more than 12 U of RBC. Transfusion data were compared with a theoretically optimal transfusion model with a fixed ratio between units of RBC, FFP, and platelets. The difference between the observed and expected amounts of blood products was calculated. RESULTS: The patients (82%) received insufficient amounts of FFP and platelets when compared to the calculated amounts. The total numbers of transfused FFP and platelets were on average 50% lower than the calculated amounts. Regression models showed an increase of FFP and platelets with increasing amounts of RBC but not in sufficient quantities. CONCLUSION: Exsanguinating trauma patients receiving massive transfusions are subject to 'blind' transfusion. This is associated with insufficient transfusion of both FFP and platelets, which may aggravate bleeding. A 'blind' transfusion strategy consisting of a validated guideline with a predefined ratio of the different blood products, timing of laboratory tests as well as a sound logistic protocol facilitating this procedure, involving the blood bank and treating physicians, is needed urgently.

Allogeneic hematopoietic cell transplantation (HCT) offers the possibility of curative therapy for patients with myelodysplastic syndromes (MDS), chronic myelomonocytic leukemia (CMML), and acute myelogenous leukemia (AML). However, post-HCT relapse remains a major problem, particularly in patients with high-risk cytogenetics and in patients who cannot tolerate consolidation chemotherapy (eg, due to previous toxicity). We assessed the toxicity and efficacy of 10-day decitabine (Dec), fludarabine (Flu), and 2 Gy total body irradiation (TBI) as a new conditioning regimen for allogeneic HCT in patients with MDS, CMML, or AML. Thirty patients were enrolled, including 11 with MDS, 2 with CMML, and 17 with AML. Patients received 20 mg/m2/day Dec on days -11 to -2, 30 mg/m2/day Flu on days -4 to -2, and 2 Gy TBI on day -1, followed by infusion of a donor stem cell graft on day 0. Postgrafting immunosuppression consisted of cyclosporin A and mycophenolate mofetil. At a median follow-up of 443 days, the overall survival was 53%, relapse incidence was 27%, and nonrelapse mortality was 27%. The incidence of severe acute (grade III/IV) graft-versus-host disease (GVHD) was 27%, and that of (predominantly mild) chronic GVHD was 60%. Immunomonitoring studies revealed that specific CD8+ T cell responses against epigenetically silenced tumor-associated antigens (TAAs), including cancer-testis antigens (MAGE-A1/A2/A3 and PRAME) and RHAMM, occurred more frequently in patients who had received Dec/Flu/TBI conditioning (8 of 11 patients) compared with a control group of patients who had received only Flu/TBI conditioning (2 of 9 patients). In summary, Dec/Flu/TBI conditioning proved feasible and effective and enhanced the induction of TAA-reactive CD8+ T cell responses in vivo, which may contribute to disease control post-transplantation.

Dendritic cell (DC)-based vaccination is an appealing strategy to boost graft-versus-tumor immunity after allogeneic stem cell transplantation (allo-SCT), and thereby prevent or counteract tumor recurrence. By exploiting minor histocompatibility antigens (MiHA) presented on hematopoietic cells, donor CD8 T-cell immunity can be selectively targeted to patient's hematological tumor cells without the risk of inducing graft-versus-host disease. Previously, we demonstrated that silencing RNA (siRNA) of programmed death-ligand 1 (PD-L1) and PD-L2 on DCs markedly augments the expansion and function of MiHA-specific CD8 T cells. However, previously applied methods based on electroporation or lipid nanoparticles were either incompatible with target antigen mRNA delivery or required complex manufacturing compliant to Good Manufacturing Practice. Here, we investigated whether transfection using lipoplexes composed of PD-L1 and PD-L2 siRNAs plus SAINT-18:DOPE (ie, SAINT-RED) is an effective and feasible clinical-grade method in DC vaccine manufacturing. We observed that a single siRNA/SAINT-RED transfection resulted in efficient and long-term knockdown of the PD-1 ligands without affecting DC maturation or viability. Furthermore, we demonstrated that SAINT-RED can be heat sterilized without loss of function, facilitating its use in aseptic DC vaccine production. Finally, we showed that the established transfection method can be combined with target antigen mRNA or peptide loading to efficiently stimulate MiHA-specific T-cell expansion and cytokine production. Together, these findings indicate that the developed PD-L siRNA/SAINT-RED transfection protocol in combination with MiHA mRNA or peptide loading can be applied in the generation of clinical-grade DC vaccines to boost antitumor immunity after allo-SCT.

Allogeneic stem cell transplantation (allo-SCT) can be a curative therapy for patients suffering from hematological malignancies. The therapeutic efficacy is based on donor-derived CD8+ T cells that recognize minor histocompatibility antigens (MiHAs) expressed by patient’s tumor cells. However, these responses are not always sufficient, and persistence and recurrence of the malignant disease are often observed. Therefore, application of additive therapy targeting hematopoietic-restricted MiHAs is essential. Adoptive transfer of MiHA-specific CD8+ T cells in combination with dendritic cell (DC) vaccination could be a promising strategy. Though effects of DC vaccination in anti-cancer therapy have been demonstrated, improvement in DC vaccination therapy is needed, as clinical responses are limited. In this study, we investigated the potency of program death ligand (PD-L) 1 and 2 silenced DC vaccines for ex vivo priming and in vivo boosting of MiHA-specific CD8+ T cell responses. Co-culturing CD8+ T cells with MiHA-loaded DCs resulted in priming and expansion of functional MiHA-specific CD8+ T cells from the naive repertoire, which was augmented upon silencing of PD-L1 and PD-L2. Furthermore, DC vaccination supported and expanded adoptively transferred antigen-specific CD8+ T cells in vivo. Importantly, the use of PD-L silenced DCs improved boosting and further expansion of ex vivo primed MiHA-specific CD8+ T cells in immunodeficient mice. In conclusion, adoptive transfer of ex vivo primed MiHA-specific CD8+ T cells in combination with PD-L silenced DC vaccination, targeting MiHAs restricted to the hematopoietic system, is an interesting approach to boost GVT immunity in allo-SCT patients and thereby prevent relapse. Electronic supplementary material The online version of this article (doi:10.1007/s00262-015-1668-6) contains supplementary material, which is available to authorized users.

In HLA-matched allogeneic hematopoietic stem cell transplantation (SCT), donor T cells can mediate graft-versus-leukemia/lymphoma (GvL) reactivity and graft-versus-host disease (GvHD) by recognition of minor histocompatibility antigens (MiHA).1–4 Only a minority of MiHA shows hematopoiesis-restricted expression, and donor T cells for these MiHA may induce beneficial GvL reactivity without GvHD. The number of well-characterized MiHA with therapeutic relevance based on hematopoiesis-restricted expression remains limited and only 25% and 40% of recipients transplanted with grafts from sibling and unrelated donors, respectively, are eligible for therapies targeting known hematopoietic MiHA.3,4 Thus, in order to increase the efficacy and applicability of cellular therapy for selective GvL induction, more hematopoiesis-restricted MiHA with balanced population frequencies in common HLA molecules must be identified. Here, we investigated the therapeutic significance of a MiHA encoded by ARHGDIB.5 We demonstrated hematopoiesis-restricted gene expression with the exception of intermediate mRNA expression in endothelial cells and showed that T cells recognized LB-ARHGDIB-1R presented by HLA-B*07:02 on primary leukemic cells, but not on [interferon-gamma (IFN-γ)]-treated fibroblasts and keratinocytes. To evaluate potential toxicity against endothelial cells, we tested T cell recognition of LB-ARHGDIB-1R on human umbilical vein endothelial cells (HUVEC) and found only limited reactivity under inflammatory conditions. Furthermore, we demonstrated in vivo targeting of LB-ARHGDIB-1R in eight out of ten patients who were screened for post-transplant specific T-cell responses. In one patient with relapsed lymphoma, high T-cell frequencies were induced after donor lymphocyte infusion (DLI), coinciding with long-lasting anti-lymphoma immunity without GvHD. Our data thus support the relevance of LB-ARHGDIB-1R as a therapeutic target with the potential to induce selective GvL reactivity.

Allo-SCT followed by DLIs can establish long-term remissions in multiple myeloma (MM) patients. In many patients, however, the immunotherapeutic graft-versus-tumor (GVT) effect is moderate and not sustained, implying that immune suppression is mediated, among other factors, by regulatory T cells (Tregs) or myeloid-derived suppressor cells (MDSCs). Towards a better understanding and, eventually, manipulation of the immune-regulatory mechanisms in transplanted MM patients, we retrospectively sought a correlation between DLI outcome and circulating CD14(+) MDSCs, CD14(-) MDSCs and Tregs in 53 MM patients before their first DLI. We found significantly elevated frequencies of highly suppressive CD14(+) MDSCs, CD14(-) MDSCs and Tregs in pre-DLI samples from patients. Higher frequencies of Tregs, but not of MDSCs, were significantly associated with non-responsiveness to DLI. Furthermore, a lower frequency of Tregs predicted the development of chronic GVHD, which, in turn, displayed a high association with GVT. Elevated Treg frequencies before DLI were also associated with significantly shorter PFS and OS. Hence, our data reinforce the idea of active suppression of antitumor responses by Tregs in MM patients and therefore suggest that targeting patient Tregs before DLI may improve outcome of DLI.

Results of Allogeneic Stem Cell Transplantation (Allo-SCT) as part of first-line therapy for multiple myeloma (MM) are conflicting.1–7 The 96 month long-term followup of the EBMT trial showed a significantly prolonged PFS and OS for Auto-/Allo-SCT as compared to double Auto-SCT, both in the intention-to-treat analysis and in the patients who actually received their allocated treatment.8 One of the conclusions from that study was that a follow-up of longer than 5 years is necessary for a correct interpretation of the value of Auto-/Allo-SCT in MM. Here we present the long-term follow-up (median 113 months) results of the donor versus no-donor (DvND) comparison of patients who were included in the HOVON-50 study.9 In this study the effect of thalidomide combined with Auto-SCT after high-dose melphalan 200 mg/m2 (HDM200) was evaluated. PFS and OS were not statistically different, neither in the donor versus no-donor comparison nor in the patients that received their allocated therapy, i.e. the Allo-SCT or maintenance (α-interferon or thalidomide, given until relapse or progression) following the Auto-SCT. Despite the extended follow-up time, there was no benefit observed for using Allo-SCT as part of first-line therapy in myeloma. Out of 536 patients randomized in the HOVON-50 study, 260 patients were eligible for the DvND analysis; 122 patients with and 138 patients without an HLA-identical donor (Figure 1). Eligibility criteria included treatment with Auto-SCT, and for inclusion in the donor group patients had to have a fully matched 10/10 sibling donor. M-protein type, ISS stage, median age and remission status were well balanced.6 The data for this update were analyzed as available on August 15th, 2014. Conditioning for the Allo-SCT was low-dose total body irradiation (TBI; 2 Gy) and GvHD prophylaxis consisted of cyclosporine and mycophenolate mofetil. The HOVON-50 study was approved by the ethics committees of the participating centers and was conducted in accordance with the Declaration of Helsinki. The trial was registered at www.trialregister.nl (NTR238; ISRCTN06413384). Figure 1. Design of the study and patient flow. The European Group for Blood and Marrow Transplant criteria were used to evaluate response. The primary endpoints were progression-free survival (PFS) and overall survival (OS) from Auto-SCT. The secondary endpoints were the impact of prognostic factors, and PFS and OS from the start of treatment for those patients who received their allocated therapy, that is, Allo-SCT or maintenance therapy with thalidomide or α-interferon (denoted herein as PFStr and OStr). For some of the endpoints the Kaplan-Meier survival curves were crossing (Figure 2) indicating a violation of the proportional hazards assumption. In that case, a standard log-rank test and Cox regression analysis are not optimal statistical methods.10 Therefore we used the so-called restricted mean survival time (RMST) method11,12 for all analyses to compare PFS and OS between donor versus no-donor, which has been implemented in Stata13 (StataCorp. 2013. Stata: Release 13. Statistical Software. College Station, TX: StataCorp LP). The difference in RMST within 10 years (RMST10y) was calculated, together with the 95% confidence interval (CI). In view of the fact that in the HOVON-50 trial patients had been randomized to induction treatment with either VAD or TAD, a treatment arm was included as a covariate. To compare our results with those previously published, we also evaluated the prognostic value of donor availability for the VAD and TAD subgroups separately, as well as in subgroups according to ISS stage (I vs. II vs. III), β-2 microglobulin (β-2M; ≤ 3 vs.> 3 mg/l) and the presence or absence of deletion 13 (determined by FISH and/or by karyotyping).14 Kaplan-Meier curves were generated to illustrate differences between subgroups. All reported P values are two-sided and have not been adjusted for multiple testing, and a significance level alpha = 0.05 was used. Figure 2. Kaplan-Meier survival curves. Actuarial rates of PFS (A) and OS (B) according to availability of an HLA-identical sibling of patients included in the HOVON-50 study. PFS and OS are presented as from the date of autologous SCT. Actuarial rates of PFStr ... The best response as determined by CR was 43% for patients with a donor and 38% for patients without a donor (P= 0.41). The 8-year and 10-year PFS were 25% and 17%, respectively, for patients with a donor and 18% and 16%, respectively, for the patients without a donor (Figure 2A). RMST10y was 6 months longer in the donor group (95% CI −5 to 16, P =0.29), which was not statistically significant. RMST10y was also not significantly different between donor and no-donor in the subgroups of VAD and TAD patients. β-2 M> 3mg/L was associated with an 11 month lower RMST10y (95% CI 0 to 22, P=0.04), but in the DvND comparison no significant difference in PFS was found for patients with low or high β-2 M. This was also apparent for ISS stage and the presence or absence of deletion 13, determined either by FISH (available for 61 % of no-donor and 57 % of donor patients) or by karyotyping (available for 82 % of no-donor and 83 % of donor patients). The 8-year and 10-year OS were 47% and 42%, respectively, for patients with a donor and 43% and 33%, respectively, for the patients without a donor (Figure 2B). RMST10y was 4 months longer in the no-donor group (95% CI -15 to 7, P=0.46). This non-significant 4.1 months longer RMST10y for OS in the no-donor patients was also observed within the subgroups of VAD and TAD patients, both P=0.6. β-2 M > 3 was associated with a 17 month lower RMST10y (95% mg CI 6 to 28, P=0.003), but the difference in RMST10y between donor and no-donor patients in each of the low or high β-2 M subgroups was less than 1 month (P=0.9). There was also no significant difference in PFS within the subgroups according to ISS and the presence or absence of deletion 13. The cumulative incidence of non-relapse mortality at 96 months after Auto-SCT was 16% in the donor group versus 3% in the no-donor group (P 3 predicted for reduced survival (P=0.02). However, no benefit for Allo-SCT as compared to maintenance was found in this patient category. Neither ISS nor deletion 13 had a significant impact on OStr. With the longest follow-up (median 113 months after Auto-SCT therapy) in published studies as yet, we found no benefit for Allo-SCT as part of first-line therapy on PFS and OS. The positive graft-versus-myeloma effect as demonstrated by the significantly reduced incidence of relapse in the donor arm did not compensate for the higher TRM (Figure 3). As in our previous analysis there was a trend for prolonged PFS in patients treated with the allocated Allo-SCT, however due to late relapses this did not lead to a significant benefit when compared to patients receiving maintenance following Auto-SCT. The absence of a benefit in our updated study may be due to better outcome for the no-donor group as compared with the Italian and EBMT studies, while the outcome with regard to PFS and OS of patients with a donor seemed rather similar. Our study was initiated later, therefore bortezomib and lenalidomide could be routinely given to patients with relapsed disease. It may be that the availability of these new anti-myeloma agents for relapse explains the comparable survival of the patients who did receive their allocated therapy, although the PFS curves diverge after 30 months in favor of the Allo-SCT group. There were 6 patients who had progressive disease at a time point of more than eight years, and we are aware that these patients were still alive between 3 and 25 months following the occurrence of the progressive disease. An important query is whether in this era of effective MM strategies, Allo-SCT should be completely abandoned or whether it could still be an option for patients with high risk features like 17p deletion or patients with an early first relapse.15 In the case that this topic is explored, alternative procedures are essential to allow effective and safe post Allo-SCT strategies to prevent both TRM and early relapse and the initiation of specific graft-versus-myeloma effects.

Contains fulltext : 57804.pdf (Publisher’s version ) (Closed access) We retrospectively analysed the outcome of voluntary unrelated donor (VUD)-SCT in 56 patients after conditioning without or with ATG. All received partially lymphocyte-depleted grafts. Four of 17 patients (24%) who were not given ATG rejected their grafts, as did one of 33 (3%) conditioned with ATG (P=0.02). The incidences of acute graft-versus-host disease grade III/IV were 29 and 6%, respectively (P=0.02), and probabilities of 1-year transplant-related mortality were 64% (95% CI, 44-84%) and 27% (95% CI, 12-42%), respectively (P=0.004). Projected at 3 years, probability of survival was 18% (95% CI, 2-34%) after conditioning without ATG and 60% (95% CI, 43-70%) after conditioning with ATG (P=0.002). Probabilities of disease-free survival (DFS) were 18% (95% CI, 2-34%) and 45% (95% CI, 27-63%), respectively (P=0.005). Patients who did not receive ATG had a probability of current DFS of 18% (95% CI, 3-34%) and this was 60% (95% CI, 43-77%) for the patients conditioned with ATG (P

Analysis of changes in recipient and donor hemopoietic cell origin is extremely useful to monitor the effect of stem cell transplantation (SCT) and sequential adoptive immunotherapy by donor lymphocyte infusions (DLI). We developed a sensitive and accurate method to quantify the percentage of recipient and donor cells by real-time PCR using single nucleotide polymorphisms (SNPs) as markers. Allele-specific PCR of seven SNPs resulted in specific markers for donor or recipient in 97% of HLA-identical sibling pairs. Both, recipient- and donor-derived hemopoietic cells can be simultaneously analyzed in 67% sibling pairs. We expect this can be increased to approximately 99% by developing three additional SNP-PCR. Serial dilution of SNP-positive DNA into either SNP-negative DNA or water revealed a detection limit of 0.1-0.01% depending on the amount of input DNA and start C(t) of the used SNP-PCR. Application of our real-time SNP-PCR method for a CML patient treated by allogeneic SCT and DLI demonstrated its feasibility to follow donor T-cell chimerism and early detection of residual and recurrent autologous hemopoiesis in response to treatment. This detailed monitoring of the genetic origin of hemopoietic cells, in particular immune effector cells and target cells after SCT and DLI, may substantially contribute to understanding of the mechanisms that play a role in the success of treatment.

Allogeneic Hematopoietic Stem Cell Transplantation Provides Cure for Adult Patients with Hemophagocytic Lymphohistiocytosis (HLH) : A Retrospective Study of The Chronic Malignancies and Inborn ErrorsWorking Parties (CMWP and IEWP) of The EBMT

Item does not contain fulltext We performed a retrospective analysis of the European Group for Blood and Marrow Transplantation database comparing the outcomes of multiple myeloma patients who received tandem autologous followed by allogeneic PSCT (auto-allo) with the outcomes of patients who underwent a reduced intensity conditioning allograft (early RIC) without prior autologous transplant. From 1996 to 2013, we identified a total of 690 patients: 517 patients were planned to receive auto-allo and 173 received an early RIC allograft without prior autologous transplant. With a median follow-up of 93 months, 5-year PFS survival was significantly better in the auto-allo group, 34% compared with 22% in the early RIC group (P

Cellular immunotherapy can be an effective adjuvant treatment for multiple myeloma (MM), as demonstrated by induction of durable remissions after allogeneic stem cell transplantation. However, anti-myeloma immunity is often hampered by suppressive mechanisms in the tumor micro-environment resulting in relapse or disease progression. To overcome this immunosuppression, new cellular immunotherapies have been developed, based on the important effector cells in anti-myeloma immunity, namely T cells and natural killer cells. These effectors can be modulated to improve their functionality, activated by dendritic cell vaccines, or combined with immune stimulating antibodies or immunomodulatory drugs to enhance their efficacy. In this review, we discuss promising pre-clinical and clinical data in the field of cellular immunotherapy in MM. In addition, we address the potential of combining these strategies with other therapies to maximize clinical effects without increasing toxicity. The reviewed therapies might pave the way to effective personalized treatments for MM patients.

Contains fulltext : 138260.pdf (Publisher’s version ) (Closed access) BACKGROUND: Outcome of Philadelphia-positive acute lymphoblastic leukaemia (Ph+ ALL) improved significantly with the introduction of tyrosine kinase inhibitors (TKIs). Autologous stem cell transplantation (ASCT) has never been considered a standard of care in this setting. The aim of our study was to analyse if results of ASCT improved in the era of TKIs. PATIENTS AND METHODS: One-hundred and seventy-seven adults with Ph+ ALL treated with ASCT in first complete remission were analysed for the impact of year of transplantation on outcome. Additional analysis was performed including 32 patients for whom detailed data on the use of TKIs and the status of minimal residual disease were collected. RESULTS: The probability of the overall survival (OS) at 3 years increased from 16% for transplants performed between 1996 and 2001 to 48% between 2002 and 2006 and 57% between 2007 and 2010 (P

Contains fulltext : 137156.pdf (Publisher’s version ) (Closed access) Application of G-CSF in AML is controversial as leukemic blasts may express receptors interacting with the cytokine, which may stimulate leukemia growth. We retrospectively analyzed the impact of G-CSF use to accelerate neutrophil recovery after auto-SCT on outcome. Adults with AML in first CR autografted between 1994 and 2010 were included. Nine hundred and seventy two patients were treated with G-CSF after auto-SCT whereas 1121 were not. BM and PB were used as a source of stem cells in 454 (22%) and 1639 (78%) cases, respectively. The incidence of relapse at 5 years in the BM-auto-SCT group was 38% for patients receiving post-transplant G-CSF and 43% for those not treated with G-CSF, P=0.46. In the PB-auto-SCT cohort, respective probabilities were 48% and 49%, P=0.49. No impact of the use of G-CSF could be demonstrated with respect to the probability of leukemia-free survival: in the BM-auto-SCT group, 51% for G-CSF(+) and 48% for G-CSF(-), P=0.73; in PB-auto-SCT group, 42% for G-CSF(+) and 43% for G-CSF(-), P=0.83. Although G-CSF administration significantly shortened the neutropenic phase, no beneficial effect was observed with regard to non-relapse mortality. In patients with AML, the use of G-CSF after auto-SCT is not associated with increased risk of relapse irrespective of the source of stem cells used.

Item does not contain fulltext In this prospective study we analyzed pre-emptive donor leukocyte infusions (DLI) in 82 consecutive patients transplanted with partially T cell-depleted grafts for acute myeloid leukemia, acute lymphoid leukemia, chronic myeloid leukemia, refractory anemia with excess of blasts, refractory anemia with excess of blasts in transformation and multiple myeloma. Donors were HLA-identical siblings. Patients without significant acute (>grade 1) and/or chronic GVHD were scheduled to be treated with DLI (35 patients) and 31 actually received DLI. Patients who developed acute GVHD >grade 1 and/or chronic GVHD were not scheduled to receive DLI and served as a comparison group (47 patients). The median interval between BMT and DLI was 22 weeks. The first six patients received 0.7 x 10(8) CD3+ cells/kg body weight (b.w.). Five out of these six patients developed acute GVHD (grade 1: n = 2, grade 3: n = 2 and grade 4: n= 1) which was more frequent and more severe than we had anticipated. In the next 25 patients the number of T lymphocytes was diminished to 0.1 x 10(8) CD3+ cells/kg b.w. which resulted in less frequent and less severe GVHD. Eight patients in this group developed acute GVHD (grade 1: n = 4, grade 2: n = 4) and three patients had limited chronic GVHD. Patients in the DLI group needed more time to establish complete donor chimerism confirmed by a higher number of mixed chimeras at 6 months after BMT. The projected 3-year probability of disease-free survival was 77% for the 35 patients intended to treat with DLI and 45% for the patients of the comparison group (P = 0.024). Relapse rate at 36 months after transplantation was 18% in the patients who were intended to treat with DLI and 44% in the comparison group (P = 0.026). We conclude that pre-emptive DLI is feasible and generates favorable relapse rates in patients who are at high risk for relapse. Furthermore, the incidence and severity of GVHD disease after DLI is dependent on the number of CD3+ cells infused.

Patients with relapsed/refractory chronic lymphocytic leukemia (CLL) have excellent responses with kinase or BCL2 inhibitors, but patients with high risk cytogenetics (del(17p) and/or del(11q)) do not seem to achieve long-term disease control. Allogeneic hematopoietic stem cell transplantation (alloHCT) can result in sustained progression-free survival. As non-relapse mortality (NRM) after alloHCT is partly age-dependent, alloHCT is preferably considered in younger high cytogenetic risk CLL patients, but data of early NRM and longer-term PFS lack for this age group. We focused in this study on younger allo-transplanted CLL patients ( Figure. Figure. Disclosures Dreger: Novartis: Speakers Bureau; Gilead: Speakers Bureau; Janssen: Consultancy; Novartis: Consultancy; Gilead: Consultancy; Roche: Consultancy. Gramatzki:Janssen: Other: Travel/Accommodation/Expenses, Research Funding. Delgado:Janssen: Consultancy, Honoraria; Novartis/GSK: Consultancy, Honoraria; Gilead: Consultancy, Honoraria; Roche: Consultancy, Honoraria, Research Funding; Infinity: Research Funding. Schoenland:Jansen: Honoraria, Other: financial support of conference participation, Research Funding; Prothena: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; GSK: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding. Schetelig:Sanofi: Honoraria.

BACKGROUND: The advent of the Bruton's tyrosine kinase inhibitor ibrutinib has improved the outlook of patients with CLL and MCL failing chemoimmunotherapy (CIT). However, the impact of ibrutinib on the feasibility and safety of a subsequent alloHCT is unknown. Here we present results of the ibrutinib cohort of an ongoing EBMT survey on the outcome of alloHCT following prior exposure to pathway inhibitors (PI) in patients with CLL or lymphoma (EBMT study code LWP 2013-N-03/CMWP 44204425). DESIGN: Eligible were patients aged >18 years registered with the EBMT data office for a planned alloHCT for CLL or lymphoma after pre-exposure to ibrutinib at any time before transplant. Baseline patient, disease, and transplant data were collected from MED-A forms. Centers were requested to provide additional treatment and follow-up information. Statistical analysis used Gray's test to assess the impact of baseline characteristics on the cumulative incidence of relapse (REL) in a competing risk framework. RESULTS: As of July 4, 2016, 38 patients (84% male) were evaluable in the ibrutinib cohort. Diagnosis was CLL in 28 patients, MCL in 9 patients, and follicular lymphoma (FL) in 1 patient. The median age was 51 (33-68) years and the median number of treatment lines prior to ibrutinib 2 (1-9). Eight of the 9 patients with MCL but none of the other patients had a prior autoHCT. Patients had been on ibrutinib for a median of 190 (39-432) days. In 2 patients, ibrutinib had been stopped because of disease progression >100d before transplant, whereas the interval between ibrutinib withdrawal and alloHCT was 15-100d in 30%, 4-14d in 51%, and 0-1d in 14% of the patients. Of the CLL patients, 43% had a TP53 lesion, and 87% and 79% met at least one of the 2007 and 2014 EBMT criteria for high-risk CLL, respectively, including PI failure in 29%. Disease status at alloHCT was sensitive in 78% of the CLL patients, and in 60% of the patients with lymphoma. Conditioning was reduced-intensity in 60% of the transplants and included in-vivo T cell depletion with ATG (71%) or alemtuzumab (11%) in the majority of cases. Donors were identical siblings in 26%, matched unrelated in 66%, and partially matched unrelated in 8%, with PBSC (89%) being the predominant stem cell source (bone marrow 8%, cord blood 3%). The median time to reach neutrophils of >0.5/nl and platelets of >20/nl was 17 (10-26) and 15 (10-46) d post transplant, respectively. Acute GVHD grade 2-4 (3-4) was observed in 37% (10%) of 30 evaluable patients, and limited and extensive chronic GVHD occurred in 24% and 16% of 25 patients at risk. With a median observation time of survivors of 8 (1-24) months, there were only 2 non-relapse deaths, translating into a 1-year non-relapse mortality (NRM) of 6% (95%CI 0-15%). 1-year REL, progression-free survival, and overall survival was 36%, 61%, and 73% for CLL, and 14%, 75%, and 75% for lymphoma. In the 25 evaluable patients with CLL, PI-sensitive compared to refractory disease status at alloHCT tended to be associated with a lower 1-y REL (29% vs 60%; p 0.071), whereas prior PI failure, TP53 status, duration of ibrutinib exposure, interval between ibrutinib withdrawal and alloHCT, and conditioning intensity had no significant impact on REL. CONCLUSIONS: Ibrutinib for bridging to alloHCT for CLL and MCL does not appear to adversely affect engraftment, GVHD risk, and NRM. Patients with CLL still responding to ibrutinib at the time of alloHCT might benefit from ibrutinib bridging as our preliminary results indicate that also after PI exposure sensitive disease translates into a lower risk of relapse. Therefore, ibrutinib may improve the perspective of CIT-refractory patients scheduled for alloHCT. The optimum timing of ibrutinib administration in the interrelation to alloHCT in CLL and MCL needs to be defined by additional studies. Disclosures Dreger: Gilead: Consultancy; Janssen: Consultancy; Novartis: Speakers Bureau; Gilead: Speakers Bureau; Novartis: Consultancy; Roche: Consultancy. Michallet:Bristol-Myers Squibb: Consultancy, Honoraria, Research Funding; Pfizer: Consultancy, Honoraria; Novartis: Consultancy, Honoraria; Pfizer: Consultancy, Honoraria; Astellas Pharma: Consultancy, Honoraria; MSD: Consultancy, Honoraria; Genzyme: Consultancy, Honoraria. Berg:Celgene: Other: Travel Funding; Astellas: Other: Travel Funding; Alexion: Other: Travel Funding. Niederwieser:Novartis Oncology Europe: Research Funding, Speakers Bureau; Amgen: Speakers Bureau. Montoto:Gilead: Research Funding; Roche: Honoraria. Schetelig:Sanofi: Honoraria.

BACKGROUND: Allo-HSCT is an accepted treatment option for patients with relapsed and refractory B cell lymphomas or high-risk chronic lymphocytic leukemia (CLL). This treatment option is currently reshaped by the introduction of pathway inhibitors (PWI) as ibrutinib, idelalisib, and venetoclax. However, the mid- and long-term efficacy and toxicity of these drugs are not yet fully defined and most patients need to discontinue their treatment over time because of disease progression or intolerance. Allo-HSCT could induce long-term disease control with curative potential especially in poor prognosis patients. In case of disease relapse after transplantation, patient's outcome is a major concern and the prognosis is dismal. In this context, PWI could be very promising in the context of lowering the bulky leukemic cells before proceeding to allo-HSCT but also when used after transplantation to treat or prevent disease relapse. The purpose of the present study is to provide information on the safety and efficacy of ibrutinib when administered after allo-HSCT for mantle cell lymphoma (MCL) or CLL. DESIGN: We included in this study adult patients who had been registered with the EBMT for an allo-HSCT for CLL or MCL and who received ibrutinib for treating disease recurrence or persistence at any time after transplant. Baseline patient, disease, and transplant data were collected from MED-A forms. Centers were requested to provide additional post-allo-HSCT treatment and follow-up information. RESULTS: A total of 30 patients, 22 (73%) males, who had undergone allo-HSCT between September 2002 and December 2015 were included in this study. Diagnosis was CLL in 27 (90%) patients and MCL in 3 patients. The median age at transplantation was 55 (range: 38-66) years and the median number of treatment lines prior to transplantation was 3 (1-8). Before transplantation, in CLL patients, del17p was present in 10/27 (37%) of cases and del11q was present in 4/27 (15%) other cases. Prior to allo-HSCT, 4 patients (2 CLL and 2 MCL) had received an auto-HSCT and 4 other CLL patients had been exposed to ibrutinib for a median of 261 (205-376) days. Disease response at allo-HSCT was sensitive in 20/27 (74%) of the CLL patients, and in 1/3 of the MCL patients. Conditioning was reduced-intensity in 20/30 (67%) of the transplants and included in-vivo T cell depletion in the majority of cases (17/20). Donors were identical siblings in 23% with PBSC being the predominant stem cell source (28/30). Acute GVHD grade II-IV was observed in 5/30 (16.6%) patients (3 grade II, 2 grade III), while chronic GVHD had occurred in 15/29 patients but had resolved in 11/15 patients prior to ibrutinib. Patients received ibrutinib for relapse after allo-HSCT (n=27, median time from allo-HSCT to progression: 22 (0.5-53) months), 1 for stable disease and 2 after positive MRD. Donor lymphocyte infusions (DLI) were administered concomitantly with ibrutinib in 3/30 patients. Ibrutinib was generally well tolerated; major toxicities observed after ibrutinib commencement were secondary cancers in 3 patients and toxic epidermiolysis in 1 patient. De novo chronic GVHD occurred in 2 patients, in one of them in the context of DLI. Overall, 23/30 (77%) patients had responsive disease after ibrutinib (33% reached CR), 21/27 with CLL (8 CR) and 2/3 with MCL (2CR). At the last follow-up, 20 (67%) patients are still on ibrutinib after a median exposure of 395 (17-703) days, 19 of them with ongoing response (8 in CR) and one too early for response assessment. Ten patients had discontinued ibrutinib, 4 because of toxicity (2 secondary cancers, 2 skin toxicity), and 6 because of disease progression. In total, 4 deaths (all CLL) were observed and were due only to progression. With altogether 8 progression events, the median PFS after start of ibrutinib of all 30 patients was 23 months, the one year OS and PFS probabilities were 92%. There was no effect of del17p or time from allo-HSCT to ibrutinib start on PFS. CONCLUSION: Ibrutinib can be safely administered for CLL/MCL relapse after allo-HSCT, with an efficacy at least similar to non-transplanted patients with high-risk disease. Figure 1 Figure 1. Disclosures Dreger: Gilead: Speakers Bureau; Janssen: Consultancy; Novartis: Speakers Bureau; Novartis: Consultancy; Gilead: Consultancy; Roche: Consultancy. Corradini:Gilead: Honoraria; Servier: Honoraria; Sanofi: Honoraria; Takeda: Honoraria; Roche: Honoraria; Novartis: Honoraria; Janssen: Honoraria; Celgene: Honoraria; Takeda: Consultancy. Schetelig:Sanofi: Honoraria.

Introduction Primary plasma cellleukemiais a rare and aggressive disorder. While autologous stem cell transplantation has been shown to improve outcome, response durations are short and further strategies are needed. Data is needed to help guide clinicians on the best approach to manage this disease. Therefore in an era of novel agents and improved autologous and allogeneic transplantation strategies this study compared different transplantation approaches in the treatment of PCL. Material and Methods A retrospective analysis was undertaken of theEuropean Group for Bone Marrow Transplantation (EBMT) experience of patients with primary PCL undergoing hematopoietic stem cell transplantation between 1998 and 2012. Only patients who had achieved complete response, partial response or stable disease prior to transplantation were included. Patients with progressive or relapsed disease were excluded. Data was collected using MED A and MED B forms. The primary end point was overall survival and data was analysed according to the information on the planned transplant strategy reported at the time of first transplant (intent-to-treat principle). Results A total of 460 patients were identified and categorised into 4 transplant groups. 1. One patient did not proceed to Allo 2. Four patients not classified The follow up period ranged from 1 to 208 months with a median follow up of 48.9 months. Patients undergoing allo upfront were found to have the worst overall survival. Compared to single auto this was statistically significant with a Hazard Ratio (HR) 1.85 (p=0.007). Compared to auto-allo the allo up front group had an even higher risk HR 3.18 (p=0.018). The double auto versus single auto had a HR 1.39 which was not statistically significant (p=0.28). The auto-allo compared to single auto has a time varying effect with a higher mortality at the beginning and better risk afterwards. The effect on average is not significant HR 0.58 (p=0.24). When we consider time varying effects three periods were examined, 0-12, 12-36 and 36-60 months. Allo upfront is still confirmed to be inferior to single auto and also to auto-allo in the first year. Auto-allo has a trend to superiority versus single or double auto in the mid-term. However due to lack of follow up data in the auto-allo group it is difficult to assess possible benefits of auto-allo in the long term as there are few patients still at risk. Our initial analysis does suggest an advantage. Preliminary analysis suggests that the auto-allo group enjoy the best progression free survival. The superiority of auto-allo versus allo upfront may be attributed to the higher rates of non-relapse related mortality observed in allo upfront. Conclusion Our preliminary work has demonstrated that allo upfront is associated with the worst overall survival. Further data is needed to reliably assess the outcome of auto-allo group which appears to be superior. Therefore we plan to request additional information from involved centres to strengthen the power of our results. Table Table. Disclosures Foà: Ariad: Speakers Bureau; Pfizer: Speakers Bureau; BMS: Consultancy; Celgene: Consultancy, Speakers Bureau; Amgen: Consultancy, Speakers Bureau; Gilead: Consultancy, Speakers Bureau; Janssen-Cilag: Consultancy, Speakers Bureau; Genetech: Consultancy; Roche: Consultancy, Speakers Bureau. Garderet:Takeda: Consultancy; Amgen: Consultancy; BMS: Consultancy, Honoraria; Novartis: Consultancy.

Fifteen consecutive patients with Philadelphia chromosome (Ph)-positive chronic myeloid leukaemia (CML) who relapsed from T-cell-depleted bone marrow transplantation (BMT) were successfully treated with donor leucocyte infusions (DLIs). Chimaerism was analysed using red blood cell phenotyping (RCP), and the results were compared with cytogenetic analysis and outcome of qualitative and quantitative polymerase chain reaction (PCR) for breakpoint molecules. In all patients, an increase in autologous erythrocytes and/or a decrease in donor red cells indicated relapse. Donor erythrocytes started to increase from 4 to 20 (median 12) weeks after DLI. At 6 and 12 months after DLI, complete donor chimaerism was found in 11 and 15 patients, respectively, and all patients were in cytogenic remission. A high percentage of autologous red cells at the time of DLI predicted pancytopenia. During relapse and after DLI, the percentage of autologous red cells was strongly correlated with the reappearance and disappearance of Ph-positive metaphases (r = 0.90; P < 0.001 and r = 0.96; P < 0.001 respectively). The same was true for the correlation between the percentage of autologous red cells and positivity/negativity in PCR for Bcr-Abl breakpoint molecules (r = 0.94; P < 0.001). A normalized Bcr-Abl dose of greater than 10-3 in real-time quantitative PCR correlated well with relapse and the presence of autologous red blood cells (r = 0.77; P < 0.001). We conclude that RCP is a sensitive, easy to perform and fast technique for the prediction of pending relapse after BMT for CML. RCP also predicts the response to DLI and the occurrence of bone marrow aplasia after DLI.

In an effort to reduce the relapse rate after unpurged autologous bone marrow transplantation (ABMT) in patients with acute myeloid leukemia (AML) in first complete remission (CR1), the standard conditioning regimens (cyclophosphamide/busulphan and cyclophosphamide/TBI) were intensified by adding idarubicin. Seventeen patients received a continuous infusion of 21 mg idarubicin/m2/day for 2 consecutive days in addition to the standard preparative regimen. Thirteen patients served as a historical control group. The 2-year disease-free survival (DFS) of 82% in the study group was significantly (P = 0.047) better compared to 46% DFS in the control group. The relapse rate (RR) was also significantly lower (7% vs 45%; P = 0.035) in the study group. The median time to reach a white cell count (WCC) of 0.5 x 10(9)/l was 20 days in the study group vs 17 days (P = NS) in the control group. The median time until recovery of the platelet counts to 20 x 10(9)/l was 152 days in the study group vs 57 days (P = NS) in the control group. The hypoplasia in the study group resulted in a trend towards a higher need for transfusions: a median number of 38 units of erythrocytes vs 23 units in the control group (P = NS) and 23 units of platelet vs 18 units in the control group (P = NS). This pilot study suggests that addition of idarubicin to the standard conditioning regimens may improve DFS and overall survival (OS) of patients with AML treated with ABMT in CR1. These results should be confirmed in a prospective randomized study.

One hundred and eighty-one consecutive patients with standard-risk leukaemia were transplanted with HLA-identical sibling grafts depleted of lymphocytes using counter-flow centrifugation. In 116 patients, standard conditioning was intensified by the addition of anthracyclines. Multivariate analysis revealed significantly more acute GVHD > or = grade 2 and a trend towards more chronic GVHD in patients conditioned with the addition of anthracyclines. For all patients the risk for chronic GVHD, but not for acute GVHD increased with a higher number of T cells in the graft. The projected 5-year probability of relapse was significantly lower in the group of patients conditioned with anthracyclines; 26% versus 52% (P = 0.015). In multivariate analysis the addition of anthracyclines to the conditioning regimen was the only significant factor contributing to a lower probability of relapse. The projected 5-year probability of leukaemia-free survival [LFS] in the patients conditioned with and without the addition of anthracyclines was 56% and 36%, respectively (P = 0.004). In multivariate analysis the addition of anthracyclines to the conditioning regimen correlated significantly with a lower number of mixed chimaeras in patients at 6 and 12 months after BMT. Mixed chimaerism at 6 months after transplantation did not significantly correlate with a higher incidence of relapse in further follow-up. In contrast, mixed chimaerism at 12 months after BMT was significantly associated with higher relapse rate. We conclude that the addition of anthracyclines to the conditioning regimen improves outcome of BMT using T-cell-depleted grafts.

Thirty-five patients with myelodysplastic syndromes (MDS) were treated with BMT between 1986 and 1994. Their median age was 41 years (range 23-60). Thirteen patients had transfusion-dependent refractory anaemia (RA). Twenty-two patients suffered from more advanced stages of MDS, 15 being in complete remission (CR) after chemotherapy. In 31 recipients, pretransplant conditioning consisted of cyclophosphamide and TBI with or without the addition of idarubucin; four patients were conditioned with other schedules. Donors were genotypically HLA-identical and MLC-negative siblings in 32, and others in three cases. All patients received a graft depleted of 98% of T lymphocytes using counterflow centrifugation. Fourteen patients are alive and in continuous remission with a median follow-up of 20 months (range 15-113) after BMT. Seven patients relapsed between 3 and 18 months after BMT and subsequently died. Fourteen transplantation-related deaths occurred. Outcome in patients under and over 40 years old was comparable. The probability of disease-free survival (DFS) at 2 years after BMT was 39% (95% confidence interval (CI), 22-56%). Considering patients with HLA-identical and MLC-negative sibling donors transplanted for RA (n = 11) or more advanced stages of MDS in CR (n = 14), the probabilities of DFS were 73% (95% CI, 47-99%) and 42% (95% CI, 15-69%), respectively. This indicates that BMT with lymphocyte-depleted grafts can cure a substantial number of relatively old patients with MDS, especially when grafts from HLA-identical and MLC-negative siblings are used and patients are suffering from RA.

Item does not contain fulltext Allogeneic stem cell transplantation (allo-SCT) can induce remission in patients with hematologic malignancies due to graft-versus-tumor (GVT) responses. This immune-mediated antitumor effect is often accompanied by detrimental graft-versus-host disease (GVHD), however. Both GVT and GVHD are mediated by minor histocompatibility antigen (MiHA)-specific T cells recognizing peptide products from polymorphic genes that differ between recipient and donor. In this study, we evaluated whether mismatches in a panel of 17 MiHAs are associated with clinical outcome after partially T cell-depleted allo-SCT. Comprehensive statistical analysis revealed that DNA mismatches for one or more autosomal-encoded MiHAs was associated with increased relapse-free survival in recipients of sibling transplants (P = .04), particularly in those with multiple myeloma (P = .02). Moreover, mismatches for the ubiquitous Y chromosome-derived MiHAs resulted in a higher incidence of acute GVHD grade III-IV (P = .004), whereas autosomal MiHA mismatches, ubiquitous or restricted to hematopoietic cells, were not associated with severe GVHD. Finally, we found considerable differences among MiHAs in their capability of inducing in vivo T cell responses using dual-color tetramer analysis of peripheral blood samples collected after allo-SCT. Importantly, detection of MiHA-specific T cell responses was associated with improved relapse-free survival in recipients of sibling transplants (P = .01). Our findings provide a rationale for further boosting GVT immunity toward autosomal MiHAs with a hematopoietic restriction to improve outcomes after HLA-matched allo-SCT.

Item does not contain fulltext No abstract available.

Allogeneic stem cell transplantation (allo-SCT) can cure hematological malignancies by inducing alloreactive T cell responses targeting minor histocompatibility antigens (MiHA) expressed on malignant cells. Despite induction of robust MiHA-specific T cell responses and long-term persistence of alloreactive memory T cells specific for the tumor, often these T cells fail to respond efficiently to tumor relapse. Previously, we demonstrated the involvement of the coinhibitory receptor programmed death-1 (PD-1) in suppressing MiHA-specific CD8+ T cell immunity. In this study, we investigated whether B and T lymphocyte attenuator (BTLA) plays a similar role in functional impairment of MiHA-specific T cells after allo-SCT. In addition to PD-1, we observed higher BTLA expression on MiHA-specific CD8+ T cells compared with that of the total population of CD8+ effector-memory T cells. In addition, BTLA’s ligand, herpes virus entry mediator (HVEM), was found constitutively expressed by myeloid leukemia, B cell lymphoma, and multiple myeloma cells. Interference with the BTLA–HVEM pathway, using a BTLA blocking Ab, augmented proliferation of BTLA+PD-1+ MiHA-specific CD8+ T cells by HVEM-expressing dendritic cells. Notably, we demonstrated that blocking of BTLA or PD-1 enhanced ex vivo proliferation of MiHA-specific CD8+ T cells in respectively 7 and 9 of 11 allo-SCT patients. Notably, in 3 of 11 patients, the effect of BTLA blockade was more prominent than that of PD-1 blockade. Furthermore, these expanded MiHA-specific CD8+ T cells competently produced effector cytokines and degranulated upon Ag reencounter. Together, these results demonstrate that BTLA–HVEM interactions impair MiHA-specific T cell functionality, providing a rationale for interfering with BTLA signaling in post-stem cell transplantation therapies.

Contains fulltext : 108338.pdf (Publisher’s version ) (Closed access) Recently, we demonstrated that reduced intensity conditioning (RIC) followed by partial T-cell-depleted SCT creates a platform for inducing the graft-versus-myeloma effect by adjuvant immunotherapy. Here, we evaluated mHA-specific T-cell responses in a multiple myeloma (MM) patient who was treated with RIC-SCT followed by donor lymphocyte infusion (DLI) and subsequent recipient DC vaccination. We isolated a mHA-specific CTL clone with the capacity to target MM tumor cells from this patient experiencing long-term CR. This CTL clone recognizes an HLA-A3-restricted mHA and mediates killing of both primary MM cells and the MM-cell line U266, while BM-derived fibroblasts are not recognized. CTL-specific T-cell receptor (TCR) transcripts could be detected by quantitative PCR analysis in both peripheral blood and BM during tumor remission. Interestingly, a strong increase of CTL-specific TCR transcripts at the BM tumor site was observed following DLI and recipient DC vaccination, while the TCR signal in peripheral blood decreased. These findings illustrate that the approach of partial T-cell-depleted RIC-SCT followed by post-transplantation immunotherapy induces mHA-specific T-cell responses targeting MM tumor cells. 01 september 2012

Introduction: Acute myeloid leukemia (AML) with 17p abnormalities (abn(17p)), usually affecting TP53 locus, carries a very poor prognosis due to high refractoriness to conventional chemotherapy, with long-term survival of less than 5%. Allogeneic stem cell transplantation (SCT) appears as the only potential curative option in high-risk AML. To specifically address outcomes after SCT in patients with abn(17), we retrospectively analysed data from the EBMT registry. Methods: De novo or secondary AML with abnormal karyotype transplanted between 2000 and 2013 have been allocated. From a dataset of 5495 patients with AML undergoing SCT, we included only those patients for whom data were sufficient to confirm the presence of abn(17p) resulting in a loss or a disruption of the TP53 locus. Results: One hundred thirty-nine patients have been selected including 125 patients (90%) in first remission (CR1) and 14 (10%) in second remission. For further analysis, we focused on the 125 patients in CR1. Median age was 54 (range, 18-69) year-old and the median follow-up was 21 (range, 3-146) months. Eighty-five percent of the patients had a de novo AML, while 15% had a secondary AML. Abn(17p) was associated to a monosomal karyotype in 83% of patients, complex karyotype in 91%, monosomy 5 or 5q deletion (-5/5q-) in 55%, monosmy 7 (-7) in 39% and both -5/5q and -7 in 27%, respectively. Median time from diagnosis to CR1 was 57 (range, 18-170) days. Fifty-one (41%) of the patients received a myeloablative conditioning regimen and 73 (59%) had a reduced-intensity conditioning regimen. The vast majority of patients (70%) had a karnofsky performance status of more than 90% at the time of SCT. The 2-year overall survival (OS) and leukemia-free survival (LFS) were 28% and 24%, respectively. The 2-year non-relapse mortality (NRM) was 15%, and 2-yr relapse incidence (RI) was 61%. The cumulative incidence of grade II to IV acute graft-versus-host disease (GvHD) was 24% and that of chronic GvHD was 21%. In multivariate analysis, the presence of a -5/5q- in addition to abn(17p) was significantly and independently associated with worse OS, LFS and higher RI. Age and donor type did not correlate with outcome. Conditioning intensity was not statistically associated with OS, LFS and NRM when adjusted for patients'age. Conclusion: In contrast to the dismal prognosis reported for AML patients harboring abn(17p) undergoing conventional chemotherapy, allogeneic SCT provides long-term responses in about 25% of a selected group of patients harboring this cytogenetic abnormality at diagnosis and transplanted in CR1. Post and pre transplant targeted therapy may further improve results. Disclosures Milpied: Celgene: Honoraria, Research Funding.

Background: Relapse is the most frequent cause of failure after alloSCT for acute leukemia (AL). Unlike in CML, infusion of donor lymphocytes (DLI) is of limited efficacy in overt hematological relapse. Hence, it may be preferable to give DLI in complete hematological remission (CHR) after alloSCT, to exploit the allogeneic graft-versus-leukemia (GvL) effect either as maintenance in high risk patients (prophylactic DLI, proDLI), or as early intervention to prevent hematological recurrence in case of decreasing donor chimerism or minimal residual disease (preemptive DLI, preDLI). However, no systematic analysis of this strategy is available so far, neither concerning the optimal way of application, nor with respect to safety and clinical efficacy. Here, the Acute Leukemia Working Party of the EBMT presents results from a registry-based survey on 343 patients with AML (n=266) or ALL (n=77), who received DLI in CHR after alloSCT. Patients: Median age was 48y, 64% of patients had received alloSCT from a matched sibling, 36% from an 8/8 matched unrelated donor. Disease status at time of alloSCT was CR1/CR2/advanced in 68%/14%/17% of cases, respectively. Patients had received standard/reduced intensity conditioning in 53%/47% of cases. Before alloSCT, 55% had received in vivo T-cell depletion (TCD), 16% ex vivo TCD, 10% in vivo plus ex vivo, and 19% no TCD. Reasons for preDLI were persisting mixed or decreasing donor cells chimerism (n=167, 49%) and persisting or recurrent minimal residual disease (MRD; n=32, 9%). ProDLI without any sign of leukemia was given to 144 patients (42%) with high risk disease Results: Median follow up from DLI1 was 6.5 years (range, 1.1-14.5). Median interval from alloSCT to first DLI (DLI1) was 180 days (range, 15-1178). Patients received a median of 2 infusions with the median CD3+ cell dose at DLI1 being 1x106/kg (range, 0.1-163). Reasons to discontinue DLI were: number of planned infusions reached (56%), GvHD (17%), disease progression (13%), and documented improvement of donor chimerism (6%). At 5y from DLI1, cumulative incidence of leukemia relapse was 43% and 28% in patients receiving preDLI for MRD and mixed chimerism, and 28% among recipients of proDLI given for maintenance in high risk disease. The corresponding 5y OS rates were 55%, 66% and 64%, 5y-LFS rates were 52%, 57% and 58%. Efficacy of preDLI could be directly demonstrated by decreasing MRD in 71% (15/21) and by improvement of donor chimerism in 68% (110/163) of informative patients. Furthermore, hematologic improvement was observed in 13 patients following proDLI. Cumulative incidences of acute GvHD grade II-IV and chronic GvHD after DLI were 13% and 32%, respectively. A multivariate model identified a history of aGvHD ≥grade II after alloSCT (p=0.009, HR 2.1, 95% CI 1.2-3.7), an interval from alloSCT to DLI 1 x 106/kg at DLI1 (p=0.024, HR 1.011, 95% CI 1.001-1.021) as risk factors for induction of GvHD after DLI in CHR. One hundred and thirty three patients (39%) had died at last follow-up, with relapse still being the most frequent cause of death (n=87). Sixteen patients (5% of the entire cohort) died from DLI-induced GvHD, and 29 patients died from other courses. In summary, in this large cohort of patients receiving DLI for AL in CHR after alloSCT, efficacy of preemptive DLI cells could be demonstrated in 69% of patients. About half of patients with MRD, and >70% of patients with mixed chimerism did not experience hematological relapse during a follow up period of >5 years, suggesting a clinically meaningful effect of preDLI in AL. GvHD was the most devastating complication, leading to death in 5% of patients. The identification of risk factors for GvHD may influence the selection of candidates for prophylactic and preemptive DLI in general, and may help to refine the use of DLI in this context with respect to cell dose and timing. Disclosures Schmid: Neovii: Consultancy; Janssen Cilag: Other: Travel grand. Bug:Celgene, Novartis: Research Funding; NordMedica, Boehringer Ingelheim, Gilead: Membership on an entity's Board of Directors or advisory committees; TEVA Oncology, Astellas: Other: Travel Grant. Tischer:Sanofi-Aventis: Other: advisory board. Esteve:Celgene: Consultancy, Honoraria; Janssen: Consultancy, Honoraria.

Contains fulltext : 95751.pdf (Publisher’s version ) (Open Access) Allogeneic stem cell transplantation (SCT) is a potentially curative treatment for patients with hematologic malignancies. Its therapeutic effect is largely dependent on recognition of minor histocompatibility antigens (MiHA) by donor-derived CD8 T cells. Therefore, monitoring of multiple MiHA-specific CD8 T cell responses may prove to be valuable for evaluating the efficacy of allogeneic SCT. In this study, we investigated the use of the combinatorial encoding MHC multimer technique to simultaneously detect MiHA-specific CD8 T cells in peripheral blood of SCT recipients. Feasibility of this approach was demonstrated by applying dual-color encoding MHC multimers for a set of 10 known MiHA. Interestingly, single staining using a fluorochrome- and Qdot-based five-color combination showed comparable results to dual-color staining for most MiHA-specific CD8 T cell responses. In addition, we determined the potential value of combinatorial encoding MHC multimers in MiHA identification. Therefore, a set of 75 candidate MiHA peptides was predicted from polymorphic genes with a hematopoietic expression profile and further selected for high and intermediate binding affinity for HLA-A2. Screening of a large cohort of SCT recipients resulted in the detection of dual-color encoded CD8 T cells following MHC multimer-based T cell enrichment and short ex vivo expansion. Interestingly, candidate MiHA-specific CD8 T cell responses for LAG3 and TLR10 derived polymorphic peptides could be confirmed by genotyping of the respective SNPs. These findings demonstrate the potency of the combinatorial MHC multimer approach in the monitoring of CD8 T cell responses to known and potential MiHA in limited amounts of peripheral blood from allogeneic SCT recipients.

Five patients with adult-onset metachromatic leukodystrophy (MLD) underwent allo-SCT. Conditioning was reduced in intensity and grafts were obtained from voluntary unrelated donors. All but one graft were depleted of T-lymphocytes. Patient age at transplantation varied from 18 to 29 (median, 27) years. Two patients rejected their graft and MLD progressed. The recipient of the unmanipulated graft converted to complete donor chimerism with normalization of arylsulphatase A (ARSA) levels. Despite ARSA normalization, he deteriorated. Another patient was a mixed chimera. Following escalated doses of donor lymphocyte infusions he converted to complete donor chimerism. His levels of ARSA correlated positively with the percentage of donor cells and MLD was not progressive. The fifth patient died after 35 days from complications associated with GVHD. We conclude that results of allo-SCT in symptomatic MLD patients are poor. However, allo-SCT may stop progression of MLD in selected patients. Bone Marrow Transplantation (2011) 46, 1071-1076; doi: 10.1038/bmt.2010.252; published online 1 November 2010

Contains fulltext : 87316.pdf (Publisher’s version ) (Closed access) Tumor relapse after human leukocyte antigen-matched allogeneic stem cell transplantation (SCT) remains a serious problem, despite the long-term presence of minor histocompatibility antigen (MiHA)-specific memory T cells. Dendritic cell (DC)-based vaccination boosting MiHA-specific T-cell immunity is an appealing strategy to prevent or counteract tumor recurrence, but improvement is necessary to increase the clinical benefit. Here, we investigated whether knockdown of programmed death ligand 1 (PD-L1) and PD-L2 on monocyte-derived DCs results in improved T-cell activation. Electroporation of single siRNA sequences into immature DCs resulted in efficient, specific, and long-lasting knockdown of PD-L1 and PD-L2 expression. PD-L knockdown DCs strongly augmented interferon-gamma and interleukin-2 production by stimulated T cells in an allogeneic mixed lymphocyte reaction, whereas no effect was observed on T-cell proliferation. Moreover, we demonstrated that PD-L gene silencing, especially combined PD-L1 and PD-L2 knockdown, resulted in improved proliferation and cytokine production of keyhole limpet hemocyanin-specific CD4(+) T cells. Most importantly, PD-L knockdown DCs showed superior potential to expand MiHA-specific CD8(+) effector and memory T cells from leukemia patients early after donor lymphocyte infusion and later during relapse. These data demonstrate that PD-L siRNA electroporated DCs are highly effective in enhancing T-cell proliferation and cytokine production, and are therefore attractive cells for improving the efficacy of DC vaccines in cancer patients.