Background and Aims The long-term safety and efficacy of allogeneic bone marrow-derived mesenchymal stromal cell [bmMSC] therapy in perianal Crohn’s disease [CD] fistulas is unknown. We aimed to provide a 4-year clinical evaluation of allogeneic bmMSC treatment of perianal CD fistulas. Methods A double-blind dose-finding study for local bmMSC therapy in 21 patients with refractory perianal fistulising Crohn’s disease was performed at the Leiden University Medical Center in 2012–2014. All patients treated with bmMSCs [1 x 107 bmMSCs cohort 1, n = 5; 3 × 107 bmMSCs cohort 2, n = 5; 9 × 107 bmMSCs cohort 3, n = 5] were invited for a 4-year evaluation. Clinical events were registered, fistula closure was evaluated, and anti-human leukocyte antigen [HLA] antibodies were assessed. Patients were also asked to undergo a pelvic magnetic resonance imaging [MRI] and rectoscopy. Results Thirteen out of 15 patients [87%] treated with bmMSCs were available for long-term follow-up. Two non-MSC related malignancies were observed. No serious adverse events thought to be related to bmMSC therapy were found. In cohort 2 [n = 4], all fistulas were closed 4 years after bmMSC therapy. In cohort 1 [n = 4] 63%, and in cohort 3 [n = 5] 43%, of the fistulas were closed, respectively. In none of the patients anti-HLA antibodies could be detected 24 weeks and 4 years after therapy. Pelvic MRI showed significantly smaller fistula tracts after 4 years. Conclusions Allogeneic bmMSC therapy for CD-associated perianal fistulas is also in the long-term a safe therapy. In bmMSC-treated patients, fistulas with closure at Week 24 were still closed after 4 years.

Burkitt lymphoma is an aggressive B cell malignancy accounting for 1–2% of all adult lymphomas. Treatment with dose-intensive, multi-agent chemotherapy is effective but associated with considerable toxicity. In this observational study, we compared real-world efficacy, toxicity, and costs of four frequently employed treatment strategies for Burkitt lymphoma: the Lymphome Malins B (LMB), the Berlin-Frankfurt-Münster (BFM), the HOVON, and the CODOX-M/IVAC regimens. We collected data from 147 adult patients treated in eight referral centers. Following central pathology assessment, 105 of these cases were accepted as Burkitt lymphoma, resulting in the following treatment groups: LMB 36 patients, BFM 19 patients, HOVON 29 patients, and CODOX-M/IVAC 21 patients (median age 39 years, range 14–74; mean duration of follow-up 47 months). There was no significant difference between age, sex ratio, disease stage, or percentage HIV-positive patients between the treatment groups. Five-year progression-free survival (69%, p = 0.966) and 5-year overall survival (69%, p = 0.981) were comparable for all treatment groups. Treatment-related toxicity was also comparable with only hepatotoxicity seen more frequently in the CODOX/M-IVAC group (p = 0.004). Costs were determined by the number of rituximab gifts and the number of inpatients days. Overall, CODOX-M/IVAC had the most beneficial profile with regards to costs, treatment duration, and percentage of patients completing planned treatment. We conclude that the four treatment protocols for Burkitt lymphoma yield nearly identical results with regards to efficacy and safety but differ in treatment duration and costs. These differences may help guide future choice of treatment.Electronic supplementary materialThe online version of this article (10.1007/s00277-017-3167-7) contains supplementary material, which is available to authorized users.

Cytotoxic T lymphocytes (CTLs) specific for hematopoietic-restricted minor histocompatibility antigens (mHags) are important reagents for adoptive immunotherapy of relapsed leukemia after allogeneic stem cell transplantation. However, expansion of these CTLs to therapeutic numbers is often hampered by the limited supply of antigen-presenting cells (APCs). Therefore, we evaluated whether cell-sized latex beads coated with HLA/mHag complexes HLA-A2/HA-1 or HLA-A2/HA-2 and recombinant CD80 and CD54 molecules can replace professional APCs. The artificial antigen-presenting constructs (aAPCs) effectively stimulated HA-1-and HA-2-specific CTL clones as shown by ligand-specific expansion, cytokine production, and maintenance of cytotoxic activity, without alteration of CTL phenotype. Furthermore, HA-1-specific polyclonal CTL lines were enriched as effi- IntroductionThe successful application of donor lymphocyte infusions for the treatment of relapsed leukemia after allogeneic stem cell transplantation illustrates the feasibility of adoptive immunotherapy of hematologic malignancies. 1 To minimize graft-versus-host disease, we earlier proposed the use of minor histocompatibility antigens (mHags) HA-1 and HA-2 as immunotherapeutic reagents. 2 HA-1 and HA-2 display hematopoietic-restricted tissue distribution and relevant expression on leukemic cells and their progenitors. [3][4][5][6] Moreover, cytotoxic T lymphocytes (CTLs) directed against these mHags do not cause graft-versus-host disease in an ex vivo skin explant model, 7 and coincide with complete remission of relapsed leukemia and multiple myeloma after HLA-matched HA-1-or HA-2-mismatched donor lymphocyte infusions. 8 HA-1-and HA-2-specific CTLs can be generated in vitro using peptide-pulsed or mHag-transduced autologous dendritic cells (DCs) as antigen-presenting cells (APCs). 9,10 However, expansion of CTLs is difficult due to the limited availability of donor-derived DCs. To date, several reports indicate effective stimulation of T cells by HLA/peptide ligands expressed on artificial antigenpresenting constructs (aAPCs) such as liposomes 11,12 or microbeads. [13][14][15][16] We developed aAPCs that can be manufactured under good manufacturing practice conditions and used to expand mHag-specific CTLs for adoptive immunotherapy. Hereto, cell-sized latex microbeads coated with HLA-A2/HA-1 or HLA-A2/HA-2 complexes, CD80, and CD54 were investigated for their potential to efficiently stimulate HA-1-and HA-2-specific CTL clones and lines while keeping their antigenspecific cytolytic properties. Study design mHag-specific CTL clones and polyclonal CTL lines, CD4 ؉ T helper cells, and DCsIn vivo and in vitro generation of mHag-specific CTL clones, polyclonal CTL lines, and DCs is documented in detail elsewhere. 9,17 CD4 ϩ T-helper cells (CD4 ϩ Th cells) were generated by culturing peripheral blood mononuclear cells (PBMCs) for 14 days in Iscove modified Dulbecco medium (IMDM) containing 10% pooled human serum (HS) and 0.5% standard Dutch diphtheria/pertussis/tetanus/po...

CTLs specific for hematopoietic system-restricted minor histocompatibility antigens (mHags) can serve as reagents for cellular adoptive immunotherapy after allogeneic stem cell transplantation (SCT). In the HLA-mismatched setting, CTLs specific for hematopoietic system-restricted mHags expressed solely by the non-self 'allo' HLA molecules could be used to treat relapse after HLA-mismatched SCT. The generation of mHag-specific allo-HLA-restricted CTLs requires antigen-presenting cells (APCs) expressing low numbers of endogenous peptides to avoid co-induction of undesired allo-HLA reactivities. In this study, we exploited viral evasion strategies to generate APCs expressing a controlled set of endogenous peptides. Herpesviruses persist lifelong following primary infection due to expression of viral gene products that hamper T-cell recognition of infected cells. The herpesvirus-derived proteins US6, ICP47 and UL49.5 down-regulate endogenous antigen presentation in human APCs via inhibition of the transporter associated with antigen processing. EBV-transformed B cell lines transduced with retroviral vectors encoding US6, ICP47 or UL49.5 exhibited a stable decrease in cell-surface HLA class I expression and were protected from lysis by mHag-specific CTLs. Exogenous addition of mHag peptide fully restored target cell recognition. UL49.5 showed the most pronounced inhibitory effect, reducing HLA class I expression and mHag-specific lysis up to 99%. UL49.5 also significantly diminished allo-HLA reactivities mediated by allo-HLA-specific CTLs. In conclusion, UL49.5 could be a powerful new tool to study and modulate endogenous antigen presentation.

Alloreactive T cells are involved in injurious graft rejection and graft-vs-host disease. However, they can also evoke beneficial responses to tumor Ags restricted by foreign MHC molecules. Manipulation of these alloreactivities requires information on the basis of T cell allorecognition. The vigorous T cell response to foreign MHC molecules may arise from peptide-independent recognition of polymorphic residues of foreign MHC molecules or peptide-specific recognition of novel peptides presented by foreign MHC molecules. We investigated CD8+ T cell allorecognition using recombinant HLA class I/peptide complexes. Peptide-specific allorecognition was examined using tetramers of HLA-A*0201 representing five peptides derived from ubiquitously expressed self-proteins that are known to bind endogenously to HLA-A*0201. Distinct subsets of CD8+ T cells specific for each HLA-A*0201/peptide combination were detected within four in vitro-stimulated T cell populations specific for foreign HLA-A*0201. Peptide-independent allorecognition was investigated using artificial Ag-presenting constructs (aAPCs) coated with CD54, CD80, and functional densities of a single HLA-A*0201/peptide combination for four different peptides. None of the four T cell populations specific for foreign HLA-A*0201 were stimulated by the aAPCs, whereas they did produce IFN-γ upon stimulation with cells naturally expressing HLA-A*0201. Thus, aAPCs did not stimulate putative peptide-independent allorestricted T cells. The results show that these alloreactive populations comprise subsets of T cells, each specific for a self-peptide presented by foreign class I molecules, with no evidence of peptide-independent components.