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This case reports on a patient with antineutrophil cytoplasmatic antibody–associated vasculitis and severe pulmonary and cutaneous involvement who received conventional therapy that failed and who was rescued by daratumumab therapy.

The pathogenesis of autoimmune complications triggered by SARS‐CoV2 has not been completely elucidated. Here, we performed an analysis of the cellular immune status, cell ratios, and monocyte populations of patients with COVID‐19 treated in the intensive care unit (ICU) (cohort 1, N = 23) and normal care unit (NCU) (cohort 2, n = 10) compared with control groups: patients treated in ICU for noninfectious reasons (cohort 3, n = 30) and patients treated in NCU for infections other than COVID‐19 (cohort 4, n = 21). Patients in cohort 1 presented significant differences in comparison with the other cohorts, including reduced frequencies of lymphocytes, reduced CD8+T‐cell count, reduced percentage of activated and intermediate monocytes and an increased B/T8 cell ratio. Over time, patients in cohort 1 who died presented with lower counts of B, T, CD4+T, CD8+T‐lymphocytes, NK cells, and activated monocytes. The B/T8 ratio was significantly lower in the group of survivors. In cohort 1, significantly higher levels of IgG1 and IgG3 were found, whereas cohort 3 presented higher levels of IgG3 compared to controls. Among many immune changes, an elevated B/T8‐cell ratio and a reduced rate of activated monocytes were mainly observed in patients with severe COVID‐19. Both parameters were associated with death in cohort 1., Patients with severe COVID‐19 present a particular set of immune changes in comparison to patients with mild disease and controls. These include consumption of certain monocyte and lymphocyte populations and an elevated B/T8 Ratio. COVID‐19 patients in general share a proinflammatory immunoglobulin profile with elevated proportions of IgG1 and/or IgG3.

Patients with cancer and autoimmune diseases are at higher risk of severe COVID-19. They may not develop protective immune responses following vaccination. We investigated patients' cellular and humoral immune response after two COVID-19 vaccine doses.Subjects were stratified into subgroups according to therapy and grade of immunosuppression at time of vaccination.Antibody titers were compared to healthy controls. 32/122 (26%) did not develop detectable antibody titers. Of these, 22 (66.6%) had active therapy. Patients showed significant lower antibody titers compared to controls (median 790 vs. 3923 AU/mL, p = 0.026). Patients with active therapy had significant lower antibody titers compared to those without (median 302 vs. 3952 U/L P 0.001). B-cell count was lower in the group without antibody titers (median 29.97 vs. 152.8; p = 0.002). 100% of patients under anti-CD20 therapy had no detectable antibody titer, followed by anti-TNF (66%), BTK inhibitors (50%), ruxolitinib (35.5%), TKI (14.2%), and lenalidomide (12.5%). Anti-CD20 therapy, ruxolitinib, BTK inhibitors, and anti-CD38 therapy presented significant lower antibody titers compared to controls.Patients undergoing therapy for cancer or autoimmune diseases are at higher risk of insufficient humoral immune response following COVID-19 vaccination. Furthermore, alterations in the B-cell compartment correlate with lower antibody titers.

Patients undergoing intensive chemotherapy are usually in need for central venous catheters (CVC). Due to contradictory study results, relation of insertion site and CVC-associated complication rate in these patients is not clear. We therefore retrospectively analyzed CVC-related data of all patients undergoing intensive chemotherapy with high risk of febrile neutropenia according to NCCN criteria, who received a CVC at our bone marrow transplantation unit between May 2016 and December 2019. In total, 210 patients received 281 CVC. CVC were placed via either the subclavian-vein (SCV, n = 58; 20%) or the internal-jugular-vein (IJV, n = 223; 80%). Median duration of CVC-lifetime and neutropenic days per CVC were comparable between the two groups (IJV vs SCV: 23 days vs 21 days (p = 0.16) and 12 days vs 11 days (p = 0.65)). Both, time to CVC removal due to local inflammation and time to central line-associated bloodstream infection was significantly shorter in patients with SCV catheters (p = 0.013 and p = 0.045). CVC placed in the IJV were associated with significantly less catheter-related infectious events compared with CVC placed in the SCV. This difference was consistent across different subgroups including 88 patients undergoing allogeneic stem cell transplantation.

Patients with diffuse large B-cell lymphoma (DLBCL) treated with the R-CHOP regime receive a high cumulative dose of prednisone. We used computer tomography-ascertained Hounsfield units (HU) as a surrogate parameter for bone mineral density (BMD) in three different locations of the L3 vertebral body at baseline and post-treatment. HU were measured in 50 patients with DLBCL of the previously published FLYER-trial which compared four cycles of R-CHOP + 2 × rituximab infusion to six cycles of R-CHOP in young, favorable DLBCL patients. In total, median loss was 26.8 HU in all patients over time. The median HU loss was significantly lower in the four cycles arm (21.3 HU vs. 41.3 HU, p = 0.023). In conclusion, young patients with DLBCL receiving R-CHOP have significant HU/BMD loss under treatment with R-CHOP. Patients receiving four cycles of R-CHOP had less HU/BMD loss than patients receiving six cycles.

The addition of rituximab to chemotherapy has substantially improved outcomes for patients with B-cell malignancies. The mechanisms of action of rituximab include activation of natural killer cells. Killer-cell immunoglobulin-like receptors (KIRs) mediate natural killer cell function through interaction with HLA. We evaluated the clinical impact of KIR-HLA genotypes on rituximab-containing therapy.For this post-hoc analysis, we used data from the RICOVER-60 trial (NCT00052936) as the discovery cohort and the CLL8 trial (NCT00281918) as the validation cohort. RICOVER-60 included patients aged 61-80 years with aggressive B-cell lymphoma treated with CHOP (cyclophosphamide, doxorubicin, vincristine, and prednisone) with or without rituximab. CLL8 included patients aged 30-81 years with chronic lymphocytic leukaemia treated with chemotherapy (fludarabine and cyclophosphamide; FC) with or without rituximab. We evaluated the KIR and HLA-C status of 519 patients with available blood samples in the RICOVER-60 trial and the KIR2DS1 and HLA-C status of 549 patients with available blood samples in the CLL8 trial, and evaluated their associations with event-free survival (RICOVER-60), progression-free survival, and overall survival (RICOVER-60 and CLL8).In the RICOVER-60 trial, 201 (39%) patients were positive for KIR2DS1, 79 (15%) were homozygous for HLA-C2, and 36 (7%) were positive for KIR2DS1 and homozygous for HLA-C2. In the CLL8 trial, 206 (38%) patients were positive for KIR2DS1, 75 (14%) were homozygous for HLA-C2, and 26 (5%) were positive for KIR2DS1 and homozygous for HLA-C2. In the RICOVER-60 trial, both KIR2DS1 and HLA-C status were identified as independent risk factors for survival. KIR2DS1 positivity, homozygosity for HLA-C2, and subsequent KIR2DS1-HLA-C status were associated with adverse clinical outcome in patients receiving rituximab-containing therapy (event-free survival for patients with KIR2DS1-HLA-C2/C2 vs all other patients, HR 2·6 [95% CI 1·4-4·7], p=0·0015; progression-free survival, 2·7 [1·5-5·1], p=0·0013; overall survival, 2·8 [1·5-5·4], p=0·0016) but not in patients receiving CHOP chemotherapy only (event-free survival, 0·9 [0·5-1·7], p=0·85; progression-free survival, 1·1 [0·6-2·0], p=0·81; overall survival, 1·2 [0·6-2·4], p=0·53). A significant interaction between KIR2DS1-HLA-C status and rituximab was observed (p=0·018 for event-free survival and p=0·034 for progression-free survival). In contrast to all other patients, those positive for KIR2DS1 and homozygous for HLA-C2 did not benefit from adding rituximab to CHOP chemotherapy (event-free survival, 1·9 [0·8-4·6], p=0·16; progression-free survival, 1·4 [0·6-3·4], p=0·48; overall survival, 1·6 [0·6-4·3], p=0·33). In the CLL8 trial, KIR2DS1-HLA-C status was confirmed as a predictive marker for benefit from rituximab therapy (p=0·024 for the interaction of KIR2DS1-HLA-C status and rituximab regarding progression-free survival). In contrast to all other patients, those positive for KIR2DS1 and homozygous for HLA-C2 did not benefit from adding rituximab to FC chemotherapy (progression-free survival, 2·1 [0·9-4·9], p=0·094; overall survival, 2·6 [0·5-12·7], p=0·21).Assessment of KIR2DS1 and HLA-C genotype might identify patients who would not benefit from rituximab, thereby allowing alternative therapies to be given. Further validation of these findings in prospective clinical trials is needed.F Hoffman La Roche.

Patients suffering from refractory aggressive B-cell lymphoma not responding to salvage chemotherapy have a dismal prognosis. CAR T-cells or allogeneic stem cell transplantation (SCT) are potentially curative approaches. However, obtaining a remission, and lowering tumor burden before consolidation seems crucial for long-term efficacy of both treatment modalities.In this retrospective analysis, we reviewed patients with chemoresistant aggressive B-cell lymphoma, defined as being refractory or progressive to at least second line salvage chemotherapy including the regimen immediately preceding autologous stem cell transplantation (ASCT), treated at 2 tertiary centers, who were eligible for intensive treatment using single agent high-dose (HD) melphalan to obtain a remission before consolidating therapy.We identified 36 patients that received single agent HD melphalan and ASCT as remission induction followed by CAR T-cells or allogeneic stem cell transplantation (SCT). Thirteen of the evaluable patients (39.4%) achieved a partial remission and 9 patients (27.73%) a complete remission, resulting in an overall response rate (ORR) of 66.7%. High remission rates were seen across all subgroups including patients with primary refractory lymphoma (ORR 58.3%), uncontrolled disease and high tumor burden as indicated by increased LDH levels (ORR 66.7% for patients with elevated LDH above 2 times upper limit of norm). 22 patients proceeded to allogeneic SCT and 5 to CAR T-cell therapy. Treatment related mortality of ASCT was 5.5% (2 patients, both due to infections). Two-year overall survival of all patients was 15.8%, primarily due to a high non-relapse mortality (45.5%) of allogeneic SCT patients treated with myeloablative conditioning chemotherapy.Single agent HD melphalan produces high remission rates in patients with chemoresistant, uncontrolled aggressive B-cell lymphoma and provides a window of opportunity for consolidation therapy.Patient with refractory/relapsed aggressive B-cell lymphoma after salvage therapy are an unmet medical need because of their very poor prognosis. In our retrospective analysis of 36 patients we showed that single agent high-dose melphalan can achieve high response rates (ORR 66.7%) even in uncontrolled disease enabling consolidation therapy e.g. with allogeneic stem cell transplantation or CAR T-cell therapy.

Hyperinflammation is frequently observed in patients with severe COVID-19. Inadequate and defective IFN type I responses against SARS-CoV-2, associated with autoantibodies in a proportion of patients, lead to severe courses of disease. In addition, hyperactive responses of the humoral immune system have been described. In the current study we investigated a possible role of neutralizing autoantibodies against anti-inflammatory mediators. Plasma from adult patients with severe and critical COVID-19 was screened by ELISA for antibodies against PGRN, IL-1-Ra, IL-10, IL-18BP, IL-22BP, IL-36-Ra, CD40, IFN-α2, IFN-γ, IFN-ω and serpinB1. Autoantibodies were characterized and the antigens were analyzed for immunogenic alterations. In a discovery cohort with severe to critical COVID-19 high titers of PGRN-autoantibodies were detected in 11 of 30 (36.7%), and of IL-1-Ra-autoantibodies in 14 of 30 (46.7%) patients. In a validation cohort of 64 patients with critical COVID-19 high-titer PGRN-Abs were detected in 25 (39%) and IL-1-Ra-Abs in 32 of 64 patients (50%). PGRN-Abs and IL-1-Ra-Abs belonged to IgM and several IgG subclasses. In separate cohorts with non-critical COVID-19, PGRN-Abs and IL-1-Ra-Abs were detected in low frequency (i.e. in < 5% of patients) and at low titers. Neither PGRN- nor IL-1-Ra-Abs were found in 40 healthy controls vaccinated against SARS-CoV-2 or 188 unvaccinated healthy controls. PGRN-Abs were not cross-reactive against SARS-CoV-2 structural proteins nor against IL-1-Ra. Plasma levels of both free PGRN and free IL-1-Ra were significantly decreased in autoantibody-positive patients compared to Ab-negative and non-COVID-19 controls. In vitro PGRN-Abs from patients functionally reduced PGRN-dependent inhibition of TNF-α signaling, and IL-1-Ra-Abs from patients reduced IL-1-Ra- or anakinra-dependent inhibition of IL-1ß signaling. The pSer81 hyperphosphorylated PGRN isoform was exclusively detected in patients with high-titer PGRN-Abs; likewise, a hyperphosphorylated IL-1-Ra isoform was only found in patients with high-titer IL-1-Ra-Abs. Thr111 was identified as the hyperphophorylated amino acid of IL-1-Ra. In longitudinally collected samples hyperphosphorylated isoforms of both PGRN and IL-1-Ra emerged transiently, and preceded the appearance of autoantibodies. In hospitalized patients, the presence of IL-1-Ra-Abs or IL-1-Ra-Abs in combination with PGRN-Abs was associated with a higher morbidity and mortality.To conclude, neutralizing autoantibodies to IL-1-Ra and PGRN occur in a significant portion of patients with critical COVID-19, with a concomitant decrease in circulating free PGRN and IL-1-Ra, indicative of a misdirected, proinflammatory autoimmune response. The break of self-tolerance is likely caused by atypical hyperphosphorylated isoforms of both antigens, whose appearances precede autoantibody induction. Our data suggest that these immunogenic secondary modifications are induced by the SARS-CoV-2-infection itself or the inflammatory environment evoked by the infection and predispose for a critical course of COVID-19.

Information regarding the curative role of allogeneic stem cell transplantation (allo-HCT) in systemic anaplastic large cell lymphoma (sALCL) is scarce. We analyzed the results of allo-HCT in patients with relapsed/refractory sALCL with special emphasis on the role of brentuximab vedotin (BV) as a bridge to allo-HCT. Forty-four patients (24 females, median age 38 years) with sALCL were included. Twenty-three patients (52%) received BV before allo-HCT; BV-treated patients were more heavily pretreated (≥3 lines of therapy in 74% vs. 38%, p = 0.04). Twenty-three patients (52%) were in complete remission (CR) at allo-HCT. Three-year nonrelapse mortality and incidence of relapse (IR) after allo-HCT were 7% and 40%, respectively. With a median follow-up of 39 (12-69) months for survivors, 3-year progression-free survival (PFS) and overall survival were 53% and 74%, respectively. Univariate analysis showed that heavily pretreated patients and those not in CR had a higher IR and a lower PFS. The use of BV before transplant did not impact on any of the outcomes. Allo-HCT is a curative therapeutic strategy in a significant proportion of patients with relapsed/refractory sALCL; BV does not seem to modify transplant-related outcomes but might be able to render more patients candidates for this curative treatment.

Vitamin D3 (25-OH-D3) deficiency impairs rituximab-dependent cellular cytotoxicity and the outcome of patients with diffuse large B-cell and follicular lymphomas (DLBCL). Since the optimum 25-OH-D3 serum levels for NK cell-mediated antibody-dependent cellular cytotoxicity (ADCC) are unknown, we determined the 25-OH-D3 serum levels associated with maximum NK cell-mediated ADCC. CD20 antibody-loaded CD20+ B-cell lymphoma cell lines were cultured with NK cells and ADCC activity was determined by lactate dehydrogenase release assays. Using a newly developed formula, pre-defined 25-OH-D3 serum levels were achieved with high individual precision over a wide range. NK cells from 20 healthy individuals killed antibody-treated CD20+ lymphoma cells in a concentration- and E:T ratio-dependent manner with obinutuzumab displaying a stronger ADCC activity than rituximab. Maximum NK-cell activity and ADCC were observed at 65 ng/ml 25-OH-D3, the middle of the normal range (30–100 ng/ml). 25-OH-D3 serum levels around this range should be the target in interventional trials aiming at improving NK cell-mediated ADCC by 25-OH-D3 substitution. Lower levels do not provide significant ADCC improvements in individuals with 25-OH-D3 deficiency or insufficiency and might result in the failure of interventions with 25-OH-D3.

INTRODUCTION: Mantle cell lymphoma (MCL) is a distinct subtype of lymphoma and is characterized by an aggressive clinical course in the majority of cases. In older patients, commonly used induction regimens are either anthracycline-containing CHOP-like regimens or bendamustine, both in combination with rituximab, followed by rituximab maintenance [Dreyling, 2017]. Data on efficacy and safety of rituximab in MCL are mainly derived from the randomized GLSG2000 trial, showing higher remission rates and longer FFS with R-CHOP vs. CHOP, but no significant differences in PFS in remission and OS due to limited statistical power [Lenz, 2005]. We now performed a long-term ITT analysis of CHOP vs. R-CHOP from all GLSG2000 patients pooled with the CHOP patients of the preceding GLSG1996 trial. METHODS: Of GLSG2000 and GLSG1996 trials, patients with advanced stage, previously untreated MCL, prospectively assigned to chemotherapy only (CHOP) or the combination with rituximab (R-CHOP), were included in this pooled analysis. Primary endpoints of this evaluation were FFS and OS, tested hierarchically. Secondary endpoints included PFS in first remission, OS after first treatment failure and rate of secondary malignancies. Exploratory subgroup analyses for FFS and OS were performed according to sex (female, male), MIPI (low, intermediate, high), Ki-67 ( RESULTS: Between 1996 and 2006, 385 MCL patients were included to receive either CHOP (n=201) or R-CHOP (n=184). Key patient characteristics were well balanced: Median age was 61 (37-86) versus 62 (35-84) years, 72 and 79% were male, median MIPI score was 5.78 (4.52-9.18, n=197) versus 5.80 (4.59-8.60 n=175), Ki67 ≥ 30% was seen in 17% (n=115) versus 17% (n=96) and blastoid morphology was seen in 8% (n=61) versus 8% (n=60) of patients treated with CHOP and R-CHOP, respectively. ASCT consolidation was done in 15% (n=31) and 21% (n=39) of CHOP and R-CHOP patients, respectively (p=0.15). Median FFS was 1.36 (95% CI: 1.18 - 1.66) vs. 2.07 (1.78 - 2.65) years, adjusted HR 0.62 (0.50 - 0.77), p Regarding long term toxicity, the rate of secondary malignancies after 10 years was 0,5 and 3,9% for hematological and 7 and 8% for non-hematological malignancies for CHOP and R-CHOP, respectively. In subgroup analyses, an efficacy of rituximab was seen across all MCL risk groups: HRs for FFS were 0.58 (95 % CI: 0.36 - 0.94) for MIPI high, 0.59 (0.41 - 0.84) for MIPI intermediate and 0.69 (0.49 - 0.97) for MIPI low (interaction p=0.80 for MIPI high vs. low; interaction p=0.50 for MIPI intermediate vs. low); aHRs for Ki67 ≥ 30% vs < 30% were 0.51 (0.25 - 1.05) vs. 0.73 (0.53 - 0.99; interaction p=0.056) and for blastoid vs. non-blastoid morphology 0.19 (0.04 - 1.03) vs. 0.77 (0.52 - 1.14; interaction p=0.19), respectively. CONCLUSION: We present mature results of a pooled MCL cohort, now demonstrating a statistically significantly prolonged OS for the combined immuno-chemotherapy confirming the current standard of care in first line treatment of MCL. Figure 1 Figure 1. Disclosures Bittenbring: Gilead: Consultancy; Pfizer: Consultancy. Hübel: Gilead: Consultancy, Speakers Bureau; Incyte: Consultancy, Speakers Bureau; Servier: Consultancy, Speakers Bureau; Celgene: Consultancy; EUSA: Consultancy, Speakers Bureau. Schmidt: Novartis: Consultancy, Honoraria, Other: Travel support; Kite/Gilead: Consultancy, Honoraria, Other: Travel support, Research Funding; Takeda: Consultancy, Other: Travel support; BMS: Consultancy, Other: Travel support; Janssen: Other: Travel support. Glass: Kite: Consultancy; Roche: Consultancy, Research Funding, Speakers Bureau; Riemser: Research Funding; Novartis: Consultancy; Helios Klinik Berlin-Buch: Current Employment; BMS: Consultancy. Hüttmann: Gilead: Honoraria; Celgene: Honoraria; Lead Discovery Center GmbH: Consultancy; Takeda: Honoraria, Membership on an entity's Board of Directors or advisory committees. Klapper: Takeda: Consultancy, Research Funding; Roche: Consultancy, Research Funding; Regeneron: Consultancy, Research Funding; Amgen: Research Funding. Hiddemann: Janssen: Research Funding; Roche: Membership on an entity's Board of Directors or advisory committees, Research Funding. Unterhalt: Roche: Research Funding. Dreyling: Genmab: Consultancy; BeiGene: Consultancy; Celgene: Consultancy, Research Funding, Speakers Bureau; Amgen: Speakers Bureau; Astra Zeneca: Consultancy, Speakers Bureau; Abbvie: Research Funding; Novartis: Consultancy, Speakers Bureau; Roche: Consultancy, Research Funding, Speakers Bureau; Incyte: Consultancy, Speakers Bureau; Janssen: Consultancy, Research Funding, Speakers Bureau; Gilead/Kite: Consultancy, Research Funding, Speakers Bureau; Bayer HealthCare Pharmaceuticals: Consultancy, Research Funding, Speakers Bureau.

Autologous hematopoietic stem cell transplantation (auto-HSCT) is the standard of care for patients with diffuse large B-cell lymphoma (DLBCL) who relapse/progress after first line chemoimmunotherapy. Long-term outcome of those who relapse after transplant is poor. We present the results of a retrospective study of 256 adult patients reported to the EBMT registry with DLBCL who relapsed after auto-HSCT performed between 2003 and 2013, and who received active salvage strategies. One hundred and fifty-four (60%) were male; median age was 53 years. Median time to relapse was 7 months, 65% relapsed during the first year. Overall response rate after salvage therapy was 46%. Median follow-up after first salvage therapy was 40 months (IQR 23-63 months). Overall survival (OS) at 3 years was 27% (95% CI 22-33). OS at 3 years of patients relapsing longer than 1 year after auto-HSCT was 41% (95% CI 31-53) compared with 20% (95% CI 14-24) in those who relapsed in less than 1 year. Eighty-two patients (32%) had a second HSCT, an allogeneic HSCT (allo-HSCT) in 69 cases, at a median time of 6.5 months after relapse. OS at 3 years after allo-HSCT was 36% (95% CI 25-51). In conclusion, the prognosis of patients with DLBCL that relapse after auto-HSCT is dismal. Patients who relapse in less than 1 year remain an unmet need, and should be considered for CAR T cell therapy or clinical trials. Patients who relapse after 1 year can be rescued with salvage therapies and a second HSCT. These results provide a benchmark to compare data of new prospective studies.

With an infection rate of 60–90%, the human cytomegalovirus (HCMV) is very common among adults but normally causes no symptoms. When T cell-mediated immunity is compromised, HCMV reactivation can lead to increased morbidity and mortality. HCMV antigens are processed and presented as peptides on the cell surface via HLA I complexes to the T cell receptor (TCR) of T cells. The generation of antibodies against HCMV peptides presented on HLA complexes (TCR-like antibodies) has been described, but is without therapeutic applications to date due to the polygenic and polymorphic nature of HLA genes. We set out to obtain antibodies specific for HLA/HCMV-peptides, covering the majority of HLA alleles present in European populations. Using phage display technology, we selected 10 Fabs, able to bind to HCMV-peptides presented in the 6 different HLA class I alleles A*0101, A*0201, A*2402, B*0702, B*0801 and B*3501. We demonstrate specific binding of all selected Fabs to HLA-typed lymphoblastoid cell lines (EBV-transformed B cells) and lymphocytes loaded with HCMV-peptides. After infection with HCMV, 4/10 tetramerized Fabs restricted to the alleles HLA-A*0101, HLA-A*0201 and HLA-B*0702 showed binding to infected primary fibroblasts. When linked to the pseudomonas exotoxin A, these Fab antibodies induce highly specific cytotoxicity in HLA matched cell lines loaded with HCMV peptides. TCR-like antibody repertoires therefore represent a promising new treatment modality for viral infections and may also have applications in the treatment of cancers. Electronic supplementary material The online version of this article (10.1007/s00262-020-02564-1) contains supplementary material, which is available to authorized users.

Background SAPHO syndrome has to be considered as a rare subtype of seronegative spondyloarthritis (SpA) showing typical manifestations with palmoplantar pustolosis and osteitis with hyperostosis; the clinical response of conventional or biological disease modifying drugs (DMARDs) in SAPHO syndrome are often disappointing. Whereas in SpA peripheral arthritis and inflammatory back pain represent the leading symptoms, the SAPHO patients often complain painful osteitis with hyperostosis in the sternal region as well as the palmoplantar pustulosis accompanied by synovitis preferentially in large joints including sacroiliitis. Recently, the detection of higher numbers of CD4+IL17+ lymphocytes in the peripheral blood of SAPHO patients has arisen the hypothesis that Th4/17 helper cells with their secretion of interleukin 17 (IL17) could be involved in the development of inflammation in SAPHO syndrome (1) Objectives Here we present an observational study of 12 SAPHO patients which were treated with the IL 17 blocking agent secukinumab. In addition, the fraction of CD4/17+ lymphocytes in peripheral blood specimen has been monitored on treatment. Methods Between January 2015 and February 2017 clinical activity of disease were measured in 37 SAPHO patients with a disease duration of 11 years (median). The disease activity were evaluated by the osteitis score and skin score (ranging from 0 to 6, assessed by physician), the HAQ score (ranging from 0 to 3, assessed by patient), and MRI of the osteitis lesion (activity score 0-3, analogously to 3 domains of OMERACT RAMRIS scoring synovitis, bone marrow edema, and erosions; assessed by radiologist). In this period of time 12 of them showed disease activity due to osteitis and/or skin disease, at which time the treatment of secukinumab at dosage of 300mg as monotherapy has been started, re-evaluated after at least 12 weeks (ranging from 12 to 18 weeks). For monitoring blood specimens were derived to assess the CD4+/IL17+ lymphocyte subpopulation using immunofluorescence technique commercial antibodies for CD4, IL17 (Boehringer Ingelheim, Germany). To analyse the differences between the disease activity scores prior to secukinumab treatment and after treatment we applied the nonparametric analysis according to Wilcoxon test. The p-value Results Secukinumab improved osteitis and palmoplantar pustolosis (osteitis score 3.8 to 2.4; p=0.007; PPP 3.4 to 2.4; p=0.071), the HAQ score was reduced from 1.25 to 1.0 (p=0.018), the MRI score from 2.08 to 1.58 (p=0.034). The reduction of at least two clinical activity parameters (osteitis, HAQ and/or MRI) after secukinumab treatment could be documented in all SAPHO patients who showed CD4+IL17+ lymphocytes in higher frequency of 0.4% of leukocytes of peripheral blood. Conclusion Secukinumab seems to control osteitis and PPP in SAPHO syndrome resulting in reduced burden of disease. High Th17 lymphocytes numbers seems to be associated with higher probability of clinical response to secukinumab. Reference: [1] Firinu D, Barca MP, Lorrai MM, Perra S, Cabras S, Muggianu E, Di Martino ML, Manconi PE, Del Giacco SR. TH17 cells are increased in the peripheral blood of patients with SAPHO syndrome. Autoimmunity. 2014Sep;47(6):389-94 Disclosure of Interests None declared

Mantle cell lymphoma (MCL) accounts for 5%–10% of all lymphomas. The disease’s genetic hallmark is the t(11; 14)(q13; q32) translocation. In younger patients, the first-line treatment is chemoimmunotherapy followed by autologous stem cell transplantation. Upon disease progression, novel and targeted agents such as the BTK inhibitor ibrutinib, the BCL-2 inhibitor venetoclax, or the combination of both are increasingly used, but even after allogeneic stem cell transplantation or CAR T-cell therapy, MCL remains incurable for most patients. Chronic antigenic stimulation of the B-cell receptor (BCR) is thought to be essential for the pathogenesis of many B-cell lymphomas. LRPAP1 has been identified as the autoantigenic BCR target in about 1/3 of all MCLs. Thus, LRPAP1 could be used to target MCL cells, however, there is currently no optimal therapeutic format to integrate LRPAP1. We have therefore integrated LRPAP1 into a concept termed BAR, for B-cell receptor antigens for reverse targeting. A bispecific BAR body was synthesized consisting of the lymphoma-BCR binding epitope of LRPAP1 and a single chain fragment targeting CD3 or CD16 to recruit/engage T or NK cells. In addition, a BAR body consisting of an IgG1 antibody and the lymphoma-BCR binding epitope of LRPAP1 replacing the variable regions was synthesized. Both BAR bodies mediated highly specific cytotoxic effects against MCL cells in a dose-dependent manner at 1–20 µg/mL. In conclusion, LRPAP1 can substitute variable antibody regions in different formats to function in a new therapeutic approach to treat MCL.

Vitamin D deficient patients suffering from diffuse large B-cell lymphoma (DLBCL) treated with R-CHOP have lower overall survival. This especially applies for patients with vitamin D levels ≤8 ng/ml (Bittenbring et al. J Clin Oncol. 2014; 32:3242-8). Vitamin D up to 65 ng/ml may be necessary for optimal efficacy of R-CHOP (Neumann et al. Cancer Immunol Immunother. 2018; 67:1709-18). To study the clinical benefit of vitamin D substitution, we established a fast-track schedule to appropriately substitute patients to these vitamin D levels. The pre-treatment baseline vitamin D level was determined in 99 patients included in the ongoing, randomized, multicenter, phase-III, open-label OPTIMAL>60 trial (NCT01478542) from November 2014 to July 2015. It was intended that patients reach a vitamin D level of 65 ng/ml. We calculated the dose needed using patient weight and baseline vitamin D: IU = 100 x ΔVitD3 x kg body weight (Van Groningen et al. Eur J Endocrinol. 2010; 162:805-11) and substitution started before treatment. Patients received 20.000 IU capsules of vitamin D, split over several days, with a daily maximum of 200.000 IU. Vitamin D level was checked again after substitution and at all restagings. Patients, who did not reach the intended vitamin D level of 65 ng/ml, received a second or third cycle of substitution. Baseline vitamin D level was 17.8±12.7 ng/ml. 14.3% of the patients reached the target value of up to 65 ng/ml after one substitution cycle with a mean dose of 386.000±137.000 IU. The average increase was 27.8 ng/ml, to a mean of 45.6±18.7 ng/ml. After the second substitution cycle with a median dose of 188.000±102.000 IU patients had vitamin D serum levels of 52.6±13.8 ng/ml. After a third substitution with 91.000±56.000 IU mean level of vitamin D was 56.0±9.6 ng/ml. By administering vitamin D in a dosage calculated by the van Groningen formula, we were able to increase the vitamin D level of our patients reliably after 2-3 cycles of substitution. After second and third substitution, a saturation effect was seen. The clinical effect of vitamin D substitution is the subject of the current study and the results are presently pending. The OPTIMAL>60 trial is supported by Amgen, Roche, Acrotech. Disclosures Poeschel: Roche:Other: Travel, Accommodations, Expenses;Amgen:Other: Travel, Accommodations, Expenses;Abbvie:Other: Travel, Accommodations, Expenses.Habersang:Rheinlandklinikum Neuss:Current Employment.Mahlberg:Janssen:Other: Travel, Accommodations, Expenses;Mutterhaus der Borromaeerinnen, Trier:Current Employment;Novartis:Honoraria, Other: Travel, Accommodations, Expenses;Labor Synlab:Ended employment in the past 24 months;Abbvie:Other: Travel, Accommodations, Expenses;Roche:Honoraria;Amgen:Honoraria, Other: Travel, Accommodations, Expenses;Merck:Honoraria, Other: Travel, Accommodations, Expenses.Keller:Janssen-Cilag:Consultancy, Other: Travel, Accommodations, Expenses;Takeda:Consultancy, Other: Travel, Accommodations, Expenses;Celgene:Consultancy, Other: Travel, Accommodations, Expenses;BMS:Consultancy, Other: Travel, Accommodations, Expenses;Roche:Consultancy, Other: Travel, Accommodations, Expenses;Hexal:Consultancy;Novartis:Consultancy;MSD:Consultancy;Pfizer:Consultancy;Astra-Zeneca:Consultancy;Pentixapharm:Consultancy.Viardot:Kite/Gilead:Honoraria, Other: advisory board;Roche:Honoraria, Other: advisory board;Amgen:Honoraria, Other: advisory board;Novartis:Honoraria, Other: advisory board.Stilgenbauer:Novartis:Consultancy, Honoraria, Other, Research Funding;Pharmacyclics:Consultancy, Honoraria, Other, Research Funding;GlaxoSmithKline:Consultancy, Honoraria, Other: travel support, Research Funding;Janssen-Cilag:Consultancy, Honoraria, Other: travel support, Research Funding;Mundipharma:Consultancy, Honoraria, Other, Research Funding;Genzyme:Consultancy, Honoraria, Other: travel support, Research Funding;Genentech:Consultancy, Honoraria, Other: travel support, Research Funding;F. Hoffmann-LaRoche:Consultancy, Honoraria, Other: travel support, Research Funding;Celgene:Consultancy, Honoraria, Other: travel support, Research Funding;Gilead:Consultancy, Honoraria, Other: travel support, Research Funding;AbbVie:Consultancy, Honoraria, Other: travel support, Research Funding;Boehringer-Ingelheim:Consultancy, Honoraria, Other: travel support, Research Funding;Amgen:Consultancy, Honoraria, Other: travel support, Research Funding.Held:Roche:Consultancy, Other: Travel, Accommodations, Expenses, Research Funding;BMS:Consultancy, Other: Travel, Accommodations, Expenses, Research Funding;MSD:Consultancy;Acrotech:Research Funding;Spectrum:Research Funding;Amgen:Research Funding.Bittenbring:Gilead:Honoraria, Other: Travel, Accommodations, Expenses;Bluebird Bio:Honoraria, Other: Travel, Accommodations, Expenses;Roche:Honoraria, Other: Travel, Accommodations, Expenses, Advisory board;Celgene:Honoraria, Other: Travel, Accommodations, Expenses, Advisory Board;Pfizer:Honoraria, Other: Travel, Accommodations, Expenses.

Previously we discovered antibodies against progranulin (PGRN-abs) in a protein array-based screening of sera from various rheumatic diseases. Here we conducted a study to evaluate the prevalence of PGRN-abs in seropositive and seronegative rheumatoid arthritis (RA).PGRN-abs were determined in the sera from 257 RA patients being seropositive for RF-IgM and/or ACPA-IgG and from 224 seronegative RA patients who were prospectively included in this study (total RA cohort n=481). All serum samples from the included participants were tested for RF-IgM as well as for ACPA-IgG, and PGRN-abs were determined using a previously described ELISA. Statistics was performed using the χ2 test for evaluating differences in clinical data; to evaluate independent statistical effects on the frequency of PGRN-abs status a logistic regression model with Wald-test was performed.PGRN-abs were detected in 25.3% from seropositive RA and in 21.0% from RF- and ACPA-negative RA resulting in a prevalence of 23.7% for both cohorts together. Comparing mean DAS28 values in the PGRN-abs positive cohort with the PGRN-abs negative cohort, the DAS28 value was significantly higher in PGRN-abs positive RA patients (3.81 vs. 3.50, p=0.038). A trend for higher frequencies of PGRN-abs in sera of RA patients with unfavourable characteristics such as erosive disease or requiring TNFi medication was observed.These data suggest that the determination of PGRN-abs in seronegative RA patients may reduce their seronegative status. Further studies are required to evaluate PGRN-abs as a potential diagnostic marker in RA.

Allogeneic hematopoietic stem cell transplantation (HCT) may result in long-term disease control in high-risk chronic lymphocytic leukemia (CLL). Recently, haploidentical HCT is gaining interest because of better outcomes with post-transplantation cyclophosphamide (PTCY). We analyzed patients with CLL who received an allogeneic HCT with a haploidentical donor and whose data were available in the EBMT registry. In total 117 patients (74% males) were included; 38% received PTCY as GVHD prophylaxis. For the whole study cohort OS at 2 and 5 yrs was 48 and 38%, respectively. PFS at 2 and 5 yrs was 38 and 31%, respectively. Cumulative incidence (CI) of NRM in the whole group at 2 and 5 years were 40 and 44%, respectively. CI of relapse at 2 and 5 yrs were 22 and 26%, respectively. All outcomes were not statistically different in patients who received PTCY compared to other types of GVHD prophylaxis. In conclusion, results of haploidentical HCT in CLL seem almost identical to those with HLA-matched donors. Thereby, haploidentical HCT is an appropriate alternative in high risk CLL patients with a transplant indication but no available HLA-matched donor. Despite the use of PTCY, the CI of relapse seems not higher than observed after HLA-matched HCT.

Paratarg-7, a frequent autoantigenic target, and all other autoantigenic targets of human paraproteins molecularly defined to date are hyperphosphorylated in the respective patients compared with healthy controls, suggesting that hyperphosphorylation of autoantigenic paraprotein targets is a general mechanism underlying the pathogenesis of these paraproteins. We now show that hyperphosphorylation of paratarg-7 occurs because of an additional phosphorylation of Ser17, which is located within the paraprotein-binding epitope. Coimmunoprecipitation identified phosphokinase C ζ (PKCζ) as the kinase responsible for the phosphorylation of most, and phosphatase 2A (PP2A) as the phosphatase responsible for the dephosphorylation of all hyperphosphorylated autoantigenic targets of paraproteins. Single-nucleotide polymorphisms (SNPs) or mutations of PKCζ and PP2A were excluded. However, PP2A was inactivated by phosphorylation of its catalytic subunit at Y307. Stimulation of T cells from healthy carriers of wild-type paratarg-7 induced a partial and transient hyperphosphorylation between days 4 and 18, which was maintained by incubation with inhibitors of PP2A, again indicating that an inactivation of PP2A is responsible for the hyperphosphorylation of autoantigenic paraprotein targets. We conclude that the genetic defect underlying the dominantly inherited hyperphosphorylation of autoantigenic paraprotein targets is not in the PP2A itself, but in genes or proteins controlling PP2A activity by phosphorylation of its catalytic subunit.

Treatment results of mantle cell lymphomas (MCL) are not satisfactory and novel therapeutic approaches are warranted. Because “shared” tumor antigens like the group of cancer testis antigens are only rarely expressed in MCL, we applied serological analysis of antigens using recombinant expression cloning (SEREX) to a complementary DNA library derived from five cases of MCL using the sera of eight patients with MCL in order to define MCL-associated antigens that are immunogenic in these patients and might be used as vaccines for patients with MCL. Five antigens were detected by SEREX. Four of the five detected antigens (hypothetical protein FLK10233, recombining binding protein suppressor, a chromosomal sequence, and interleukin-1 receptor associated kinase) are also expressed by a wide spectrum of normal human cells, excluding their use as vaccines. In contrast, the expression of CD52, which was detected by antibodies in the serum of an MCL patient, is restricted to hematopoietic cells. Interestingly, anti-CD52 antibodies were detected in this patient before and >2 years after allogeneic transplantation, indicating that both the autologous as well as the allogeneic immune system recognized CD52. Since the anti-CD52 monoclonal antibody alemtuzumab has shown activity in MCL, a vaccine consisting of recombinant CD52 alone or combined with passive immunotherapy using alemtuzumab warrants furthers clinical and immunological evaluation in MCL.

Lymphoplasmacytic lymphoma (LPL) is an indolent lymphoma with moderate sensitivity to conventional chemotherapy. This study investigated whether the addition of rituximab to standard chemotherapy improves treatment outcome in LPL and the subgroup of LPL patients fulfilling the criteria of Waldenstroem's macroglobulinemia (WM). A total of 69 patients with previously untreated LPL were enrolled into the trial; 64 patients were evaluable for treatment outcome. In all, 48 of the 64 LPL patients fulfilled the criteria of WM. Patients were randomly assigned to R-CHOP (rituximab, cyclophosphamide, doxorubicin, vincristine and prednisone, n=34) or CHOP (n=30). R-CHOP resulted in significantly higher overall response (OR) rate (94 vs 67%, P=0.0085) in the LPL patients and in the WM subgroup (91 vs 60%, P=0.0188). With a median observation time of 42 months, R-CHOP induced a significantly longer time to treatment failure (TTF) with a median of 63 months for R-CHOP vs 22 months in the CHOP arm in the LPL patients (P=0.0033) and in the WM subgroup (P=0.0241). There was no major difference of treatment-associated toxicity between both treatment groups. These data indicate that the addition of rituximab to front-line chemotherapy improves treatment outcome in patients with LPL or WM.

Introduction. Allogeneic hematopoietic cell transplant (HCT) is an established treatment modality that is potentially curative for many patients with acute myeloid leukemia (AML). The development of reduced intensity conditioning (RIC) allows performing HCT in elderly and/or in heavily pretreated patients and in those with comorbidities precluding the use of standard myeloablative conditioning. Post-transplant relapse remains a challenge after RIC, particularly in patients with adverse prognosis factors.The so-called "sequential" transplant approach (e.g. FLAMSA regimen combining both intensive chemotherapy and RIC HCT within the same procedure) initially developed in patients with refractory AML, could be a promising strategy to improve disease control and decrease the risk of relapse in high-risk AML patients in complete remission (CR). Patients and methods. In the current study we analyzed transplantation outcomes in a cohort of 411 adults AML patients in CR at time of transplant, treated between 2002 and 2013. Patients received a "sequential" conditioning regimen based on Fludarabine 30 mg/m2/d, high-dose aracytine 1-2 g/m2/d, amsacrine 100 mg/m2/d for 5 days and after a 3 days rest, total body irradiation (TBI) 4Gy, cyclophosphamide 50-120 mg/kg, and antithymocyte globulin (ATG) for 2 to 3 days (TBI group, n=269 [65%]). In 142 (35%) patients, TBI was substituted by IV Busulfan 3.2 mg/kg/d for 2 days, or orally equivalent dose (Bu group). 323 patients (79%) had de-novo AML and 88 (21%) had a secondary AML (with prior myelodysplastic syndrome). At time of transplant, 300 (73%) patients were in CR1 and 111 (27%) in CR2. Cytogenetic study in de novo AML was favorable in 19 patients (6%), intermediate in 102 (32%) and poor in 41 (13%). Cytogenetic data were missing in 161 (50%). 104 (25%) patients received matched related donors (MRD) and 307 (75%) unrelated donor (URD) HCT. Majority of patients (94%) received mobilized peripheral blood stem cells graft. Results. Median follow-up of surviving patients was 28 months and median age at transplant was 54 years (18-76). ANC>500/μL was achieved at a median of 17 (range, 9-74) days after HCT. Sixteen patients (4%) failed to engraft. Two year cumulative incidence of relapse (RI) and non-relapse mortality (NRM) were 22% (95%CI, 18-26%) and 22% (95%CI, 18-27%), respectively. The Kaplan-Meier estimate of overall (OS) and leukemia-free survival (LFS) at 2 years were 59% (95%CI, 54-65%) and 56% (95%CI, 50-61%), respectively. Acute GVHD (grade II-IV) occurred in 109 (28%) patients. The 2-year cumulative incidence of chronic GVHD was 31% (95%CI, 26-36), extensive in 17% (95%CI, 12-21). Two years RI, NRM, LFS and OS in TBI vs. Bu patients were 21.8% vs 21.7% (p=0.69), 29.4% vs 18.3% (p=0.008), 48.8% vs 59.6% (p=0.045) and 51.2% vs 64.0% (p=0.013), respectively. In multivariate analysis adjusted for variable with different distribution between Bu and TBI groups, the type of conditioning (TBI vs Bu) has no impact on RI, NRM, LFS and OS. Age over 55 at transplant was an independent adverse prognostic factor in multivariate analysis for NRM (hazard ratio (HR: 1.61, 95% CI: 1.00-2.61, p=0.05)), LFS (HR: 1.39, 95% CI: 1.00-1.92, p=0.05) and OS (HR: 1.55, 95% CI: 1.11-2.18, p=0.01). Being treated in an experienced center (defined as having including 10 or more transplants in the study) was associated with a significant lower RI (HR: 0.84, 95% CI: 0.75-0.93, p=0.001) and better LFS (HR: 0.91, 95% CI: 0.84-0.98, p=0.01) and OS (HR: 0.91, 95% CI: 0.84-0.98, p=0.02). Finally, transplantation from an URD was associated with a significant increase in NRM (HR: 2.11, 95% CI: 1.14-3.91, p=0.02). Of note, CR1 vs. CR2 and de novo vs. secondary AML had no impact on patients' outcome. Conclusions. These results in a rather large cohort of patients with AML suggest that a FLAMSA "sequential" regimen provided an efficient disease control in high-risk AML patients including in CR2 and secondary AML. Furthermore Busulfan and TBI based FLAMSA "sequential" regimens provide a similar outcome. These results should be confirmed in a multicenter well design randomized study. Disclosures Off Label Use: off-label drug use: antithymocyte globulin (ATG) for allo-SCT conditioning. Tischer:Sanofi-Aventis: Other: advisory board. Schmid:Neovii: Consultancy; Janssen Cilag: Other: Travel grand. Mayer:Janssen: Research Funding. Hallek:Pharmacyclics: Honoraria, Other: Speakers Bureau and/or Advisory Boards, Research Funding; Janssen: Honoraria, Other: Speakers Bureau and/or Advisory Boards, Research Funding; Boehringher Ingelheim: Honoraria, Other: Speakers Bureau and/or Advisory Boards; Mundipharma: Honoraria, Other: Speakers Bureau and/or Advisory Boards, Research Funding; Celgene: Honoraria, Other: Speakers Bureau and/or Advisory Boards, Research Funding; Gilead: Honoraria, Other: Speakers Bureau and/or Advisory Boards, Research Funding; Roche: Honoraria, Other: Speakers Bureau and/or Advisory Boards, Research Funding; AbbVie: Honoraria, Other: Speakers Bureau and/or Advisory Boards, Research Funding.

Background: Refractory or early relapsed ( Methods: At our institution we identified all patients with aggressive B-cell lymphoma (diffuse-large B-cell lymphoma and blastoid mantle cell lymphoma) who were refractory or progressive to salvage chemotherapy with R-DHAP and who had peripheral blood stem cells (> 2x10^6 CD34+/kg body weight) collected after the 1st or 2nd cycle. These patients were eligible and planned to receive salvage high-dose melphalane and autologous stem cell transplantation as remission induction before allogeneic stem cell transplantation. Allogeneic transplantation was performed from sibling donors or matched unrelated or mismatched unrelated donors: The conditioning regimen with fludarabine, busulfane, cyclophosphamide and anti-thymocyte globuline as published (Glass et al; Lancet Oncology 2014 Jun;15(7):757-66) was used in all but 2 patients. Statistical analysis was performed with Prism©; GraphPad Software; (LaJolla CA) Results: From2006 to 2015 we identified 28 patients (21 male , 7 female, age 18-67 years, median age 47 years), 25 had diffuse large B-cell lymphoma and 3 had blastoid mantle cell lymphoma. 22 had relapsed within 1 year after primary diagnosis and 6 within a median of 25.3 months. After high-dose melphalane and autologous stem cell transplantation 13 patients had a partial and 6 a complete remission. 1 patient died due to neutropenic infection, 2 patients died due to progressive disease leading to a transplant related mortality of 3.5 %. Median progression-free survival after autologous transplantation was 4.6 months. 24 proceeded to allogeneic stem cell transplantation. 8 patients had a matched related sibling, 9 had a matched unrelated donor and 7 had a mismatched unrelated donor. Transplant related mortality was 42 % in this heavily pretreated population. 2-year overall survival of all patients intended for treatment is 21 %. One of these patients with relapsed mediastinal lymphoma after allogeneic transplantation was cured by salvage radiotherapy and is in long-term remission (> 2 years). Conclusions: Salvage high-dose melphalane and autologous peripheral blood stem cell transplantation for diffuse large B-cell lymphoma as a bridge to allogeneic transplantation is potentially curative for a minor fraction of these patients. However, the remission rate of 79 % (46% PR, 21% CR) and progression-free survival of 4.6 months after high-dose melphalane and autologous stem cell transplantation provides a window of opportunity to use new drugs and cellular therapies in these poor prognosis patients. Disclosures No relevant conflicts of interest to declare.

PURPOSE of this analysis was to provide 10-year follow-up of the GCLLSG CLL3X trial which aimed at evaluating reduced-intensity conditioning (RIC) HSCT in patients with poor-risk CLL. DESIGN: The CLL3X trial included 100 patients (median age 53 (27-65) years), of whom 90 patients were allografted with blood stem cells from related (40%) or unrelated donors (60%) using fludarabine-alkylator-based RIC regimens. 24% had refractory CLL at HSCT, and 37% had a TP53 deletion and/or mutation. The 6-year follow-up of the trial including the observation that genetic risk factors such as TP53 lesions and SF3B1 and NOTCH1 mutations had no prognostic impact has been previously reported (Blood 2013;119:4851). Survival and relapse information was requested for all patients who underwent HSCT within the CLL3X trial in 9 German centres (the Canadian centre was unavailable for follow-up) and were alive at the 6-year follow-up. RESULTS: Follow-up information was received for 33/44 patients (75%) alive at the 6-year follow-up. Of these, 2 patients had experienced non-relapse mortality (NRM; 1 chronic GVHD, 1 secondary cancer) and another 2 had CLL recurrence 9.3 and 9.7 years post HSCT, respectively. With a median follow-up of survivors of 9.3 (0.6-15.5) years, 10-year NRM, relapse incidence (REL) event-free survival (EFS), and overall survival (OS) of all 90 patients allografted was 26%, 57%, 32%, and 51%, respectively. Absence of minimal residual disease (MRD) at the 12-month landmark post HSCT was highly predictive for an increased relapse risk (10-year REL 25% vs 80% if MRD was present at the 12-month landmark, p CONCLUSIONS: Long-term observation of patients allografted in the CLL3X trial confirms that RIC HSCT can provide GVL-mediated sustained disease control in a sizable proportion of patients with poor-risk CLL independent of the TP53 status. Patients who are in MRD-negative remission one year after HSCT have a 75% probability of remaining disease-free at least for 10 years. However, late relapses do occur but may benefit from strategies involving innovative pathway inhibitors. Disclosures Krämer: Gilead: Other: Travel grants; MSD: Honoraria. Stilgenbauer:AbbVie: Consultancy, Honoraria, Other: Travel grants, Research Funding; Hoffmann-La Roche: Consultancy, Honoraria, Other: Travel grants , Research Funding; Janssen: Consultancy, Honoraria, Other: Travel grants , Research Funding; Novartis: Consultancy, Honoraria, Other: Travel grants , Research Funding; Genzyme: Consultancy, Honoraria, Other: Travel grants , Research Funding; Pharmacyclics: Consultancy, Honoraria, Other: Travel grants , Research Funding; Boehringer Ingelheim: Consultancy, Honoraria, Other: Travel grants , Research Funding; Amgen: Consultancy, Honoraria, Other: Travel grants, Research Funding; Genentech: Consultancy, Honoraria, Other: Travel grants , Research Funding; Gilead: Consultancy, Honoraria, Other: Travel grants , Research Funding; Mundipharma: Consultancy, Honoraria, Other: Travel grants , Research Funding; GSK: Consultancy, Honoraria, Other: Travel grants , Research Funding; Celgene: Consultancy, Honoraria, Other: Travel grants , Research Funding; Sanofi: Consultancy, Honoraria, Other: Travel grants , Research Funding. Böttcher:Hoffmann-LaRoche: Honoraria, Other: Travel grants, Research Funding; Celgene: Research Funding; AbbVie: Honoraria, Research Funding. Ritgen:Roche: Membership on an entity's Board of Directors or advisory committees, Research Funding. Hegenbart:Jansen Cilag: Honoraria, Other: financial support of conference participation. Hallek:F. Hoffmann-LaRoche: Consultancy, Honoraria, Other: travel support, Research Funding, Speakers Bureau; Amgen: Consultancy, Honoraria, Other: travel support, Research Funding, Speakers Bureau; Janssen-Cilag: Consultancy, Honoraria, Other: travel support, Research Funding, Speakers Bureau; Celgene: Consultancy, Honoraria, Other: travel support, Research Funding, Speakers Bureau; Gilead: Consultancy, Honoraria, Other: travel support, Research Funding, Speakers Bureau; Mundipharma: Consultancy, Honoraria, Other: travel support, Research Funding, Speakers Bureau; AbbVie: Consultancy, Honoraria, Other: travel support, Research Funding, Speakers Bureau. Kneba:Amgen: Research Funding; Gilead: Consultancy, Honoraria, Other: Travel grants, Research Funding; Glaxo-SmithKline: Other: Travel grants; Roche: Consultancy, Honoraria, Other: Travel grants, Research Funding; AbbVie: Consultancy, Honoraria, Other: Travel grants; Janssen-Cilag: Consultancy, Honoraria, Other: Travel grants. Dreger:Gilead: Consultancy; Novartis: Consultancy; Gilead: Speakers Bureau; Novartis: Speakers Bureau; Janssen: Consultancy; Roche: Consultancy.

Introduction: Allogeneic hematopoietic stem cell transplantation (HCT) is a treatment for CLL that can give long disease control. Even with the availability of kinase and BCL2 inhibitors, HCT is still performed in fit patients (pts) with high-risk CLL. Almost exclusively, outcomes on matched related and unrelated donor transplantations in CLL have been published. Recently, mismatched related donors are gaining interest because of the better outcome of haploidentical HCT with post-transplantation cyclophosphamide (PTCY). Methods: All pts with CLL who received a first allogeneic HCT with a mismatched related donor and whose data were available in the EBMT registry were analyzed. Median values and ranges are reported for continuous variables and percentages for categorical variables. The probabilities of overall survival (OS) and progression-free survival (PFS) were calculated using the Kaplan-Meier method and the log-rank test for univariate comparisons. Relapse/progression and nonrelapse mortality (NRM) were analyzed together in a competing risk framework. Statistical analyses were performed using SPSS and R. Results: One-hundred-seventeen pts with CLL (74% males) underwent a mismatched related donor transplantation between 1984 and 2015 (1984-1999: 10, 2000-2004: 18, 2005-2009: 23, 2010-2016: 66). Median follow-up after HCT was 8 months (range 0-187 months). Median age at transplantation was 54 years (yrs) (range 27-71 yrs). Median time from diagnosis to HCT was 67 months (range 4-207 months). Eighteen pts (17%) had previously undergone autologous stem cell transplantation (ASCT). Disease status at HCT was CR in 16% of pts, PR in 39% and SD/PD in 45%. The Karnofsky score was known for 98 pts; 96% had a score of 70% or more at the time of HCT. Fifty-eight percent of pts received reduced-intensity conditioning, 42% myeloablative conditioning. Peripheral blood stem cells were used in 68% of pts, bone marrow in 32%. The HCT was sex matched in 41% of recipient-donor pairs. The relationship of the donor to the patient was known for 34 pts; in 53% the donor was a child, in 38% a sibling and in 6% a parent. Forty pts (38%) received PTCY as GVHD prophylaxis. In the other 77 pts various methods of T-cell depletion (TCD) were used, but not all methods were specified. At least 56% of those pts had in vivo TCD. For the whole cohort of pts OS at 2 and 5 yrs was 46% and 37%, respectively. PFS at 2 and 5 yrs was 38% and 30%, respectively. The use of PTCY did not have a significant impact on OS (49% vs. 42% at 2 yrs, 44% vs. 33% at 5 yrs, p=0.35) and PFS (45% vs. 31% at 2 yrs, 40% vs. 22% at 5 yrs, p=0.15). CI of NRM in the whole group at 2 and 5 yrs were 41% and 45%, respectively. CI of relapse at 2 and 5 yrs were 21% and 25%, respectively. The CI of NRM and relapse at 2 and 5 yrs were not statistically different in pts who received PTCY compared to other types of TCD (NRM: 38% vs. 45% at 2 yrs, 43% vs. 49% at 5 yrs, p=0.45; relapse: 17% vs. 25% at 2 yrs, 17% vs. 29% at 5 yrs, p=0.33). For the whole cohort, the incidence of acute graft-versus-host disease (aGVHD) at 100 days was 34% for grade II-IV and 16% for grade III-IV with a median time of onset of 23 days (range 4-57 days). Conclusions: Mismatched related donor HCT resulted in a 5-year PFS in 30% of the pts. This result seems only slightly inferior to matched donor transplant (5 yrs PFS 37%1). NRM was higher than expected in this cohort, but comparable to other studies on haploSCT with in vivo T-cell depleted grafts. In conclusion, a mismatched related donor HCT may be considered for high-risk chemoimmunotherapy-refractory or 17p deleted/TP53 mutated CLL pts without options for kinase and BCL2 inhibitor therapy. More data are needed to assess the value of PTCY for GVHD prophylaxis in this specific context. References: 1. Schetelig J, de Wreede L, Moreno C, et al. Risk factors for adverse outcome in patients with Chronic Lymphocytic Leukemia (CLL) undergoing Allogeneic Hematopoietic Cell transplantation (alloSCT): a Retrospective EBMT Analysis. Abstract WP024, EBMT meeting 2015. Figure 1 Figure 1. Disclosures Ciceri: MolMed SpA: Consultancy. 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. Hallek:Mundipharma: Consultancy, Honoraria, Other: travel support, Research Funding, Speakers Bureau; F. Hoffmann-LaRoche: Consultancy, Honoraria, Other: travel support, Research Funding, Speakers Bureau; AbbVie: Consultancy, Honoraria, Other: travel support, Research Funding, Speakers Bureau; Janssen-Cilag: Consultancy, Honoraria, Other: travel support, Research Funding, Speakers Bureau; Celgene: Consultancy, Honoraria, Other: travel support, Research Funding, Speakers Bureau; Gilead: Consultancy, Honoraria, Other: travel support, Research Funding, Speakers Bureau; Amgen: Consultancy, Honoraria, Other: travel support, Research Funding, Speakers Bureau. Schetelig:Sanofi: Honoraria. Kröger:Sanofi: Honoraria, Research Funding; Neovii: Honoraria, Research Funding; Riemser: Honoraria, Research Funding; Novartis: Honoraria, Research Funding.

In addition to antimicrobial activity, macrophages regulate tissue development, remodeling, and repair. In terms of neoplastic growth, tumor-associated macrophages (TAMs) are even considered pro-tumorigenic based on their angiogenic and T-cell suppressive properties. In multiple myeloma (MM) macrophages represent an abundant component of the stromal cell compartment and are believed to support proliferation, survival, and drug resistance of MM cells. Those pro-tumorigenic functions are partly instructed by T-helper (TH)-2 cytokines such as interleukin-4 (IL-4) and IL-10, which skew TAM differentiation towards an alternatively activated, or so-called M2-like, phenotype. In contrast, M1 macrophages generally considered as potent effectors in response to microbial products or interferon-γ (IFN-γ), are characterized by superior antigen-presentation, abundant production of pro-inflammatory cytokines such as interleukin-12 (IL-12), and consequently, promote a polarized type I immune response against infections as well as malignant cells. Transcriptional regulation is a key determinant for macrophage polarization. The Ikaros (IKZF1) transcription factor is critical for lymphoid development and is found in all hematopoietic progenitors as well as in T-, B-, NK-cells and macrophages. Interestingly, IKZF1 is overexpressed in MM and selectively degraded by lenalidomide, which is approved in MM therapy. The potential role of IKZF1 in modulating macrophage polarization has not been elucidated yet. Here, we show that IKZF1expression is found highly elevated in M2-like macrophages and in MM TAMs. IKZF1 deletion in human macrophages by small interfering RNA (siRNA) or by lenalidomide yields an upregulation of M1-specific cytokines (IL-12 and IL-1b), chemokines (CXCL10 and CCL5), and costimulatory molecules (CD86 and CD40) and leads to a potent TH1-TH17 response. In fact, lenalidomide-pretreated macrophages display strong tumoricidal effects when co-cultured with MM cell lines as opposed to their untreated counterparts promoting MM proliferation and viability. Utilizing immunoprecipitation-sequencing (ChIP-Seq) we reveal that IKZF1 governs IRF4 and IRF5 expression in human macrophages. Recent studies demonstrate that IRF4 controls M2 macrophage polarization, while IRF5 regulates the M1 phenotype respectively. We could show that a lenalidomid-mediated M1-phenotype induction is efficiently abrogated by IRF4 overexpression or IRF5 silencing. Overall, these findings unravel a novel role for IKZF1 in orchestrating macrophage polarization via the IRF4/IRF5 pathway. Disclosures No relevant conflicts of interest to declare.

Molecules which are exclusively or preferentially expressed by malignant cells are potential targets for immunotherapeutic approaches to the treatment of cancer.We therefore tested serum samples of 95 patients with aggressive B-cell lymphomas for antibodies against lymphoma-associated molecules using SEREX for antibodies against BCL-2, BCL-XL, MCL-1, survivin, BCL-6, CD20, LM02, PDE4B, cyclin-D2, actinin-alpha1, SCYA3, PKCbeta2, E2IG3, and HGAL.One patient had low-titered (1:100) antibodies against PKCbeta2. Antibodies against BCL-2 were detected in the sera of five patients (5.3%), with responses found more frequently in follicular lymphoma grade 3 (3/26 or 11.5%) than in diffuse large B-cell lymphomas (2/67 or 3%). Anti-BCL-2 antibodies were of the IgG class and titers ranged from 1:1,000 to 1:100,000 with highest titers before treatment and decreasing slowly during remission.The spontaneous immunogenicity of BCL-2 makes this molecule a prime candidate for vaccine approaches in BCL-2 positive B-cell lymphomas.

Multiple myeloma (MM) is considered a chronic and incurable disease due to its highly complex and heterogeneous molecular abnormalities and the support from myeloma microenvironment factors. Macrophages are an abundant component of the stromal cell compartment and are believed to support proliferation, survival, and drug resistance of MM cells. Conversely, macrophages are key immune effector cells for the therapeutic effect of monoclonal antibodies and can directly eliminate tumor cells. However, myeloma-associated macrophages (MAMs) regularly fail to exert direct effector functions. Given their abundance in MM, an attractive therapeutic approach would be to stimulate their tumoricidal activity in order to promote antitumor immunity. Lenalidomide, an immunomodulatory agent that enhances antibody dependent cell mediated cytotoxicity (ADCC), has the potential to synergize with MOR202, an anti-CD38 monoclonal IgG1 antibody currently in phase I/IIa for the treatment of MM. Furthermore, vitamin D plays a key role in regulating effector functions of human macrophages. This is closely linked to the expression of the vitamin D-1-hydroxylase CYP27B1, which catalyzes the conversion of 25-hydroxy-vitamin D (25D) to the bioactive form 1,25-di-hydroxy-vitamin D (1,25D). We have previously shown, that vitamin D promotes tumoricidal activity of macrophages and improves the efficacy of rituximab-dependent cytotoxicity (Bruns et al., Sci. Transl. Med., 2015; Bittenbring et al., JCO, 2014). Therefore, we hypothesized that the combination of MOR202 with lenalidomide and MOR202 with 1,25D would enhance the MOR202- dependent macrophage-mediated effector functions against myeloma cells. Here we report that MAMs exhibit an altered vitamin D metabolism with a reduced expression of the vitamin D receptor (VDR) and CYP27B1, while the expression of CYP24A1, which catabolizes 1,25D, is increased. As a consequence MAMs cannot convert 25D into bioactive 1,25D. Given the importance of the vitamin D pathway for antibody mediated cytotoxicity, we screened several drugs for their ability to restore the vitamin D pathway in human macrophages. We found, by RNA-sequencing, that lenalidomide treatment modulates the phenotype of monocyte-derived macrophages and isolated MAMs, and that lenalidomide significantly increases the expression of the VDR, CYP27B1 and CAMP (cathelicidin antimicrobial peptide) in these macrophages. Furthermore, we demonstrate that isolated MAMs regularly fail to eliminate primary myeloma cells, and that the lack of effector functions can be overcome by treatment with lenalidomide and vitamin D. Moreover, we show that MOR202-dependent elimination of myeloma cells is enhanced by pre-treatment of isolated MAMs with lenalidomide and supplementation of vitamin D. In summary, these data show that the bioactive form of vitamin D (1,25D) is essential for the effector functions of MAMs and that the therapeutic activation of the vitamin D pathway by lenalidomide may restore their tumoricidal effector mechanisms. Furthermore, these findings highlight that stimulating the tumoricidal activity of MAMs, an abundant component of the stromal cell compartment, with lenalidomide and 1,25D is an attractive therapeutic approach in combination with the anti-CD38 monoclonal IgG1 antibody MOR202. Disclosures Bruns: Morphosys: Research Funding; Celgene: Research Funding.

Background: Vitamin D3 deficiency impairs the rituximab-mediated antibody-dependent cellular cytotoxicity (ADCC) and the outcome of elderly patients with diffuse large B-cell lymphoma (Bittenbring et al., J Clin Oncol. 2014 Oct 10; 32(29):3242-8) treated with R-CHOP and of patients with follicular lymphoma (Kelly et al., J Clin Oncol. 2015 May 1; 33(1):1482-90. The aim of this study was to determine the optimal 25-OH-vitamin D3 (VD3) level for rituximab- and obinutuzumab-mediated ADCC. Methods: Ten individuals (5 males, 5 females; mean age: 67.7 years, range 41-79) without malignant disease or immunosuppression were included in this study after written informed consent. PBMC were isolated by density gradient centrifugation. CD16+ NK cells were separated by depleting all non-NK-cells magnetically. ADCC activity of the NK cells was tested against CD20-expressing Daudi cells without or after anti-CD20-antibody treatment with rituximab and obinutuzumab, respectively. Cytotoxic activity was assessed by LDH release by the target cells. NK cells were studied at four different VD3 serum levels: 1 (insufficient supply: < 20 ng/ml); 2 (lower normal range; 30 ng/ml), 3 (mid normal range; 65 ng/ml), and 4 (high normal range; 90 ng/ml). To achieve these levels the probands were substituted with cholecalciferol. Results: The median VD3 serum level before substitution was 10 ng/ml. The only formula for achieving predefined VD3 serum levels published by van Groningen et al. (Eur J Endocrinol. 2010; 162:805-11) failed to achieve the predetermined VD3 levels by far. Modifying this formula by using a multiplication factor of 1.8-2.2 we achieved the preset dose levels with high precision with median levels of 32.6 ng/ml, 66.4 ng/ml and 92.3 ng/ml aiming at dose levels of 30, 65 and 90 ng/ml, respectively. 2/10 test persons had starting VD3 levels above 30 ng/ml, but 6 of the remaining 8 individuals showed a significantly increased ADCC after VD3 substitution to level 2 (ranging from + 10 % to + 147 % increased Daudi lysis, p < 0.05). Further substitution to 65 ng/ml significantly increased ADCC activity in all 10 persons compared to the ADCC at 30 ng/ml (ranging from + 18.4 % to + 89.2 % increased Daudi lysis, p < 0.05). 8/10 individuals were further substituted to achieve 90 ng/ml. However, in 7/8 of these probands the NK-cell-mediated ADCC was significantly reduced compared to their values at 65 ng/ml (ranging from - 23.1 % to - 58.1 % decreased Daudi lysis, p < 0.05). The extent of the substitution-induced ADCC improvement varied individually and depended on the antibody concentration used to treat the target cells. Rituximab-mediated ADCC was significantly more affected by VD3 levels than obinutuzumab. Conclusion: Our study demonstrates for the first time that the ADCC of NK cells is optimal at median VD3 serum levels around 65 ng/ml. Our data strongly argue for a rapid vitamin D3 substitution before/at the beginning of R-CHOP treatment using the modified van Groningen formula. 65 ng/ml was chosen as the target VD3 level in the ongoing OPTIMAL>60 study of the German High-Grade Non-Hodgkin Lymphoma Study Group (DSHNHL) in elderly patients with DLBCL. This study is supported by the Eva Mayr-Stihl-Stiftung (Waiblingen, Germany). Disclosures Pfreundschuh: Roche, Janssen, Celgene: Honoraria, Research Funding.

Infiltration by macrophages represents a characteristic morphological hallmark in high grade lymphatic malignancies such as Burkitt’s lymphoma (BL). Although macrophages can in principle target neoplastic cells and mediate antibody-dependent cytotoxicity (ADCC), tumor associated macrophages (TAMs) regularly fail to exert direct cytotoxic functions. The underlying mechanisms responsible for this observation remain unclear. Here we demonstrate that inflammatory M1 macrophages kill proliferating high grade B-cell lymphoma cells by releasing the antimicrobial peptide cathelicidin in a vitamin D-dependent fashion. Moreover, we show that cathelicidin directly induces cell death by targeting mitochondria of BL-cells. In contrast, anti-inflammatory M2 macrophages and M2-like TAMs in BL exhibit an altered vitamin D metabolism resulting in a reduced production of cathelicidin and consequently in inability to lyse BL cells. However, treatment of M2 macrophages with the bioactive form of vitamin D, 1,25D, or a vitamin D receptor agonist effectively induces cathelicidin production and triggers tumoricidal activity against BL cells. Furthermore, rituximab-mediated cytotoxicity of M2 macrophages is cathelicidin dependent and vitamin D treatment of 25D deficient volunteers improves rituximab-mediated ADCC against BL cells. These data indicate that activation of the vitamin D signaling pathway may activate antitumor activity of TAMs and improve the efficacy of ADCC. Disclosures No relevant conflicts of interest to declare.

Background To investigate the impact and underlying mechanisms of vitamin-D-deficiency (VDD) on outcome of elderly (61 to 80 year-old) DLBCL patients. Methods Pretreatment 25-OH-vitamin-D serum levels from 359 patients treated in the prospective multicenter RICOVER-60 trial with 6 or 8 cycles of CHOP-14 with and without 8 cycles rituximab and 63 patients in the RICOVER-noRT study treated with 6xCHOP-14 + 8xR were determined determined by LIASION®, a commercially available chemoluminescent immunoassay. Results RICOVER-60 patients with VDD (defined as serum levels ≤8 ng/m l) and treated with rituximab had a 3-year event-free survival of 59% compared to 79% in patients with >8 ng/ml; 3-year overall survival was 70% and 82%, respectively. These differences were significant in a multivariable analysis adjusting for IPI risk factors with a hazard ratio of 2.1 [p=0.008] for event-free survival and 1.9 [p=0.040] for overall survival. In patients treated without rituximab 3-year EFS was not significantly different in patients with vitamin-D levels ≤8 and >8 ng/ml (HR 1.2; p=0.388). These results were confirmed in an independent validation set of 63 patients treated within the RICOVER-noRT study. Rituximab-mediated cellular toxicity (RMCC) against the CD20+ cell line Daudi as determined by LDH release assay increased significantly (p Conclusions VDD is a significant risk factor for elderly DLBCL patients treated with rituximab. Our results show that VDD impairs RMCC and that RMCC can be improved by vitamin-D substitution. This together with the differential effect of VDD in patients treated with and without rituximab suggests that vitamin-D substitution might result in a better outcome of these patients when treated with CHOP plus rituximab. Supported by a grant from Deutsche Krebshilfe. Disclosures: No relevant conflicts of interest to declare.

8569 Background: Vitamin D deficiency was shown to be is associated with a worse outcome in patients with non-Hodgkin's lymphoma (Drake et al., 2010) To study whether this observation could be confirmed in patients with aggressive B-cell lymphomas treated uniformly within a prospective trial, we analyzed 25-OH vitamin D serum levels in patients treated within the RICOVER-60 trial of the DSHNHL. Methods: 25-OH Vitamin D serum levels were determined with a commercial chemoluminescence immunoassay in the serum from elderly patients of the RICOVER-60 trial which compared 6 or 8 cycles of CHOP, both with and without rituximab. Results: 193 of 359 pts (53.8%) had vitamin D deficiency (8 ng/ml; the respective figures for 3-year overall survival were 70% and 82%, respectively. In R-CHOP pts these differences were significant in a multivariable analysis adjusting for IPI risk factors with a hazard ratio (HR) of 2.1 (p=0.008) for EFS and a HR of 1.9 (p=0.040) for OS. In pts treated without R effects of vitamin D deficiency were significant only for OS (HR 1.8; p=0.025), but not with respect to EFS (HR 1.2; p=0.388). These results were confirmed in an independent validation set of 63 patients treated within the prospective RICOVER-noRx study. Conclusions: Vitamin D deficiency is a significant risk factor for patients with aggressive B-cell lymphomas treated with R-CHOP. The stronger adverse effect of vitamin D deficiency in patients receiving rituximab suggests that vitamin D deficiency interferes with the R mechanisms of this antibody. A prospective study evaluating the effects of vitamin D substitution on outcome of patients receiving R-CHOP is warranted. Supported by Deutsche Krebshilfe.

Background: Minor histocompatibility antigens are antigenic peptides derived from normal cellular molecules which are presented in the context of major histocompatibility antigens (MHC class I and MHC class II). After an allogeneic hematopoietic stem cell transplantation (aHSCT) recipient-derived mHags can be recognized by T-cells of the transplanted immune system an mediate both a graft-versus-host disease and graft-versus- leukemia reaction. All mHags known to date have been identified by allo-reactive T-cells. Due to complex logistics needed for the establishment of such T-cell clones only a few mHags have been molecularly defined to date. Since patients with GvH develop antibodies against mHags and the presence of mHag antibodies has been shown to correlate with GvH and maintenance of remission [Miklos et al. (2005) Blood 105:2973-9] we set out to identify new mHags using SEREX the serological identification of antigens using recombinant expression cloning [Sahin et al. (1995) PNAS 92:11810-3]. Methods: Fibroblasts obtained by skin biopsies from patients with chronic GvH were propagated in vivo and used as a source to establish a cDNA library. cDNA was expressed by lambda phage in E.coli and expressed clones were screened for reactivity with antibodies in the serum of patients with chronic GvH. Positive clones were monoclonized and sequenced. The sequences were compared with known sequences using BLAST data bank. Reactive sera were compared with patient’s pre transplant serum and the serum from the donor to exclude pre-existing antibodies against (auto-)antigens expressed by the fibroblasts. Only neo-antigens recognized by the transplanted immune system were analyzed further. Results: cDNA libraries obtained from 7 patients with chronic GvH after HLA-identical stem cell transplantation were screened with the sera of these 7 patients by SEREX. Antibodies from one patient with chronic GvH were absent in the donor and the patients pre-transplant serum, reacted with c1ORF107. This constellation proves that the c1ORF107 antibodies in the patient’s serum developed after transplant and recognized c1ORF as a neo-antigen. The immunogenic c1ORF107 of the patient differed at previously described polymorphic position 275 (rs585627) with G in the recipient leading to glutamate and C in the donor leading to glutamine. The definition of the c1ORF107 epitopes eliciting a T-cell response by “reverse T-cell immunology” is currently underway. Conclusion: This study proves the principle that the analysis of the humoral immune response in patients with chronic GvH by SEREX allows for the definition of new mHags. SEREX in combination and followed by reverse T-cell immunology is a straight-forward approach which will expand considerably the number of molecularly defined mHags causing GvH in patients after allogeneic stem cell transplantation. Supported by Jose Carreras Foundation Germany

Background SNP309 T/G (rs2279744) causes higher levels of MDM2, the most important negative regulator of the p53 tumor suppressor. SNP72 G/C (rs1042522) gives rise to a p53 protein with a greatly reduced capacity to induce apoptosis. Both polymorphisms have been implicated in cancer. The SNP309 G-allele has recently been reported to accelerate diffuse large B-cell lymphoma (DLBCL) formation in pre-menopausal women and suggested to constitute a genetic basis for estrogen affecting human tumorigenesis. Here we asked whether SNP309 and SNP72 are associated with DLBCL in women and are correlated with age of onset, diagnosis, or patient's survival. Methods SNP309 and SNP72 were PCR-genotyped in a case-control study that included 512 controls and 311 patients diagnosed with aggressive NHL. Of these, 205 were diagnosed with DLBCL. Results The age of onset was similar in men and women. The control and patients group showed similar SNP309 and SNP72 genotype frequencies. Importantly and in contrast to the previous findings, similar genotype frequencies were observed in female patients diagnosed by 51 years of age and those diagnosed later. Specifically, 3/20 female DLBCL patients diagnosed by 51 years of age were homozygous for SNP309 G and 2/20 DLBCL females in that age group were homozygous for SNP72 C. Neither SNP309 nor SNP72 had a significant influence on event-free and overall survival in multivariate analyses. Conclusion In contrast to the previous study on Ashkenazi Jewish Caucasians, DLBCL in pre-menopausal women of central European Caucasian ethnicity was not associated with SNP309 G. Neither SNP309 nor SNP72 seem to be correlated with age of onset, diagnosis, or survival of patients.