[Interventional endoscopy in hepatico-pancreatico-biliary disease]. / Moderne Behandlungsoptionen an Gallenwegen und Pankreas in der interventionellen Endoskopie.

Interventional endoscopy in hepatico-pancreatico-biliary disease Abstract. Interventional endoscopy has undergone an exciting evolution in recent years, especially in the treatment of hepatico-pancreatico-biliary disease. However, novel endoscopes have also improved diagnostics through direct visualization of the biliary tract and the pancreatic duct, including targeted biopsy-sampling of suspicious endoluminal lesions. The use of cholangioscopy also allows for lithotripsy of complex gallstones under direct visualization. In patients with malignant biliary obstruction, endosonography(EUS)-guided insertion of novel lumen-apposing metal stents allow for extraanatomical access to intra- or extrahepatic bile ducts, if traditional transpapillary stent insertion fails. Especially in patients with altered anatomy e. g., after surgery, EUS-guided transgastric drainage may be the most sensible approach. Using similar techniques, EUS-guided drainage of the pancreatic duct is also possible. Local tumor therapy may be achieved with novel endoscopic retrograde cholangiography-guided catheter-based radiofrequency ablation and photodynamic therapy. In chronic pancreatitis, similar to biliary disease, methods of endoluminal diagnosis, lithotripsy, and transgastric drainage may also be applied. This article provides an overview of novel endoscopic techniques and methods that are available in hepatico-pancreatico-biliary disease.

Potential of immunotherapies in the mediation of antileukemic responses for patients with acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS) - With a focus on Dendritic cells of leukemic origin (DCleu).

New (non-immunotherapeutic) treatment-strategies for AML/MDS-patients are under development. Dendritic cells (DCs) and 'leukemia-derived DC' (DCleu) connect the innate and the adaptive immunesystem and (re-)activate it, in their capacity as professional antigen-presenting cells (APCs). They can be generated ex vivo from peripheral blood mononuclear cells (PBMNCs) or whole blood (WB), containing the -physiological-cellular/soluble microenvironment of individual patients using various DC/DCleu-generating methods or (for WB) minimalized 'Kits', containing granulocyte-macrophage-colony-stimulating-factor (GM-CSF) and a second response-modifier. Proof for DC/DCleu-mediated activation of the immune-system after T-cell-enriched mixed lymphocyte culture (MLC) is done by flowcytometry, demonstrating increased fractions of certain activated, leukemia-specific or antileukemic cell-subsets of the innate and the adaptive immune-system. Generation of DC/DCleu is possible independent of patients' age, MHC-, mutation- or transplantation-status. In vivo-treatment of AML-/MDS-patients with blast-modulating, DC/DCleu- inducing 'Kits' could contribute to create migratory DCs, as well as antileukemically reactivated and memory-mediating immune-cells, which patrol tissue and blood and could contribute to stabilizing disease or remissions.

Role of Interferon (IFN)α in "Cocktails" for the Generation of (Leukemia-derived) Dendritic Cells (DCleu) From Blasts in Blood From Patients (pts) With Acute Myeloid Leukemia (AML) and the Induction of Antileukemic Reactions.

Strategies to stabilize remissions by specific elimination of residual acute myeloid leukemia (AML) blasts are needed. Leukemia-derived dendritic cell (DCleu/DC) generated from myeloid blasts improve antileukemic T-cell reactivity and install T-cell memory. Interferon (IFN)α-DC methods produce DCleu from chronic myeloid leukemia-patients (pts') blood. Various INFα-containing versus other DC methods were studied to produce DCleu (evaluated by flowcytometry) from AML-pts' blast-containing mononuclear (MNC) or whole blood (WB). After DCleu/DC stimulation in mixed lymphocyte cultures, T cells' potential to gain antileukemic cytotoxicity was studied and correlated with different DC methods and DCleu/DC counts. (1) Generation of DCleu/DC: (a) "IFN-GIT" [containing granulocyte macrophage-colony stimulating factor (GM-CSF)+IFNα+ tumor necrosis factor (TNF)-α] produced DC successfully (≥10% DC, ≥5% DCleu/cells) from AML-MNC (WB) in 54 (56%), "MCM-Mimic" in 76 (75%), "Picibanil" in 83 (64%), and "Calcium-ionophore" in 42 (67%) of cases. Proportions of DC subtypes in MNC (WB) were comparable with all DC methods, (b) IFNα combinations containing only GM-CSF+IFNα or only IFNα showed low efficiency to produce DCleu/DC from MNC (WB) compared with "IFN-GIT." (2) Antileukemic functionality: DCleu/DC-stimulated T cells showed improved leukemia cytotoxicity compared with blast cells or unstimulated T cells. The highest blast proliferation (=insufficient T cells) was seen with "IFN-GIT" DC-stimulated T cells. Probability to respond to immunotherapy or to obtain blast lysis of DC-stimulated T cells correlated with high proportions of DCleu/DC after DC culture, independent of DC-generating methods. (3) Cytokine release profiles: levels of interleukin-6, IFN-γ, and interleukin-2 were significantly lower in DC culture supernatants (from MNC/WB) with "IFN-GIT" compared with "MCM," "Pici," and "Ca" DC supernatants. Our data show that (1) WB culture simulates AML-pts' in vivo situation, (2) DC generation is possible from AML-MNC (WB) with IFNα-containing and other DC methods, (3) successful IFNα-DC generation needs GM-CSF+IFNα+TNF-α (IFN-GIT); however, "IFN-GIT" produces less DCleu/DC compared with other (non-IFNα) DC methods, (4) T cells stimulated with "IFN-GIT"-produced DCleu/DC yielded comparable antileukemic cytotoxicity; however, in cases without achieved blast lysis, an increased blast proliferation was observed.

Diagnose und Monitoring bei chronisch-entzündlicher Darmerkrankung.

Diagnosis and Monitoring of Inflammatory Bowel Disease Abstract. The diagnosis and monitoring of inflammatory bowel disease (IBD) is based on several factors: Clinical history, physical examination, laboratory values (blood and stool), endoscopy, histology and imaging. No single feature establishes the diagnosis alone. In recent years, therapeutic goals in IBD have evolved from clinical endpoints to endoscopic or even histologic targets. However, these targets, e. g. mucosal healing, still have to be uniformly defined. Repeated endoscopies are ill-tolerated by patients, therefore surrogate-markers of mucosal inflammation such as calprotectin, have been investigated and showed good correlation with endoscopic findings. In Crohn's Disease, directing therapy with tight control, based, among others, on fecal calprotectin, has been superior to conventional therapy-monitoring and decision making. However, these initial results need further confirmation. Therapeutic drug monitoring (TDM) has emerged as a second monitoring entity in the long term treatment of IBD patients. Especially with the increasing use of biologics, reactive TDM (in patients who relapse) and to a lesser extent proactive TDM (in patients who are in clinical remission / stable disease) have been studied and results have been adopted in current guidelines. Future studies will have to better define treatment targets and further investigate the impact of tight-monitoring on disease outcome.

Paramunity-inducing Factors (PINDs) in dendritic cell (DC) cultures lead to impaired antileukemic functionality of DC-stimulated T-cells.

INTRODUCTION: Paramunity-inducing-Factors (PINDs) consist of attenuated/inactivated viruses of various poxvirus-genera, used in veterinary medicine as non-antigen-specific, non-immunising stimulators of the innate immune system against infectious and malignant diseases. Their danger-signaling-interactions were tested for their capacity to improve leukemic antigen-presentation on DC generated from AML-patients' blasts ('DCleu') and DC-stimulation/activation of antileukemic T-cells. METHODS: We analyzed, whether the addition of PINDs during DC cultures (15 healthy, 22 leukemic donors) and mixed lymphocyte culture (MLC, n = 15) with autologous (n = 6), allogeneic (n = 2) or T-cells after stem cell transplantation (SCT; n = 7) would alter the quality and quantity of DC, the composition of T-cell-subsets, and/or their antileukemic functionality (AF) as studied by FACS and functional Fluorolysis-cytotoxicity-assays. RESULTS: Effects on 1. DC-cultures: PINDs in DC-cultures lead to increased proportions of mature DC and DCleu, but reduced proportions of viable and overall, as well as TLR4- and TLR9-expressing DC. 2. MLC: PINDs increased early (CD8+) T-cell activation (CD69+), but reduced proportions of effector-T-cells after MLC 3. AF: Presence of PINDs in DC- and MLC-cultures reduced T-cells' as well as innate cells' antileukemic functionality. 4. Cytokine-release profile: Supernatants from PIND-treated DC- and MLC-cultures resembled an inhibitory microenvironment, correlating with impaired blast lysis. CONCLUSIONS: Our data shows that addition of PINDs to DC-cultures and MLC result in a "blast-protective-capacity" leading to impaired AF, likely due to changes in the composition of T-/innate effector cells and the induction of an inhibitory microenvironment. PINDs might be promising in treating infectious diseases, but cannot be recommended for the treatment of AML-patients due to their inhibitory influence on antileukemic functionality.

Profiles of activation, differentiation-markers, or ß-integrins on T cells contribute to predict T cells' antileukemic responses after stimulation with leukemia-derived dendritic cells.

Stem cell transplantations and donor lymphocyte infusions are promising immunotherapies to cure acute myeloid leukemia (AML). Leukemia-derived dendritic cells are known to improve antileukemic functionality of T cells. We evaluated the composition and development of distinct T-cell subtypes in AML patients (n=12) compared with healthy probands (n=5) before and during stimulation with leukemia-derived dendritic cells-containing DC (DC) or blast-containing mononuclear cells (MNC) in 0-7 days mixed lymphocyte cultures (MLC) by flow cytometry. AML patients' T-cell subgroups were correlated with antileukemic functionality before and after DC/MNC stimulation by functional fluorolysis assays. (1) Unstimulated T cells from AML patients presented with significantly lower proportions of activated, Tcm, CD137, and ß-integrin T cells, and significantly higher proportions of Tnaive and Teff compared with healthy probands. (2) After 7 days of DC or MNC stimulation, T-cell profiles were characterized by (significantly) increased proportions of activated T cells with effector function and significantly decreased proportions of ß-integrin T cells. (3) Antileukemic cytotoxicity was achieved in 40% of T cells after MNC stimulation compared with 64% after DC stimulation. Antileukemic activity after DC stimulation but not after MNC stimulation correlated with higher proportions of Tcm and Tnaive before stimulation, as well as with significantly higher proportions of activated and ß-integrin T cells. Furthermore, cutoff values for defined T-cell activation/differentiation markers and ß-integrin T cells could be defined, allowing a prediction of antileukemic reactivity. We could demonstrate the potential of the composition of unstimulated/DC-stimulated T cells for the lysis of AML blasts. Especially, AML patients with high numbers of Tnaive and Tcm could benefit from DC stimulation; proportions of activated and ß-integrin T cells correlated with increased antileukemic functionality and could serve to predict T cells' reactivity during stimulation. Refined analyses in the context of responses to immunotherapies are required.

Quality of T-cells after stimulation with leukemia-derived dendritic cells (DC) from patients with acute myeloid leukemia (AML) or myeloid dysplastic syndrome (MDS) is predictive for their leukemia cytotoxic potential.

Myeloid leukemic cells can differentiate into leukemia-derived dendritic cells (DC(leu)), presenting known/unknown leukemic-antigens. Induced anti-leukemic T-cell-responses are variable. To further elicit DC/DC(leu)-induced T-cell-response-patterns we performed (functional)flow-cytometry/fluorolysis-assays before/after mixed lymphocyte cultures (MLC) of matched (allogeneic) donor-T-cells (n=6), T-cells prepared at relapse after stem cell transplantation (n=4) or (autologous) patients'-T-cells (n=7) with blast-containing-mononuclear-cells ('MNC') or DC(leu)-containing DC ('DC'). Compared to 'MNC' 'DC' were better mediators of anti-leukaemic T-cell-activity, although not in every case effective. We could define cut-off proportions of mature DC, DC(leu), proliferating, CD4(+), CD8(+) and non-naive T-cells after 'MNC'- or 'DC'-stimulation, that were predictive for an anti-leukemic-activity of stimulated T-cells as well as a response to immunotherapy. Interestingly especially ratios >1 of CD4:CD8 or CD45RO:CD45RA T-cells were predictive for anti-leukemic function after DC-stimulation. In summary the composition and quality of DC and T-cells after a MLC-stimulating-phase is predictive for a successful ex-vivo and in-vivo anti-leukemic response, especially with respect to proportions of proliferating, CD4(+) and CD45RO(+) T-cells. Successful cytotoxicity and the development of a T-cell-memory after 'DC'-stimulation could be predictive for the clinical course of the disease and may pave the way to develop adoptive immunotherapy, especially for patients at relapse after SCT.