Crosstalk of human mesenchymal stromal cells with the cellular components of the immune system

Using the potential of immune regulatory cell populations for c ellular therapy constitutes an attractive tool to obliterate imbalances of immune responses in inflammatory disorde rs. In this context, adoptive transfer of m esenchymal stromal cells (MSCs) represents a relatively novel approach and its impact on the immune system has not been completely clarified. In this thesis we aimed to study the effects of MSCs on key immune cell types, which led us amongst others to investigate regulatory T - cell s (T Regs ), and myeloid cells. We show that MSC s utilize the anti - oxidative, immune regulatory enzyme hemeoxygenase - 1 (HO - 1) for suppressing T - cell activation directly and for inducing T Re gs (=indirect T - cell suppression) . An i nflammatory milieu generated by alloreactive T - cell s led to the so - called ‘licensing’ of the MSCs boosting their regulatory capacity. Interestingly, HO - 1 expression was substantially diminished during this process and its functions were taken over by other (up - regulated) molecules such as cyclooxygenase - 2 thereby highlighting (functional) MSC plasticity. Most MSC - based trials lack a systemic immune monitoring, which is key for interpreting the in vivo effects of MSCs. Performing a comprehensive flow cytometry - based immune screening in patients with acute graft - versus - host disease (aGVHD) , treated with either third - party MSC or placebo infusions (in a double - blinded fashion) , we were - most importantly - able to further corroborate the notion that MSCs function in vivo partly by promoting T Reg - subsets . Thereby, our data underscores the need for accompanying extens ive immune analyses to better comprehend such “bench - to - bedside” approaches. Accordingly, we carried out thoro ugh , laboratory investigations when we were the first to apply MSCs in a patient with treatment - refractory hemophagocytic lymphohistocytosis . Upon MSC infusion we could observe an increase of the immune modulating cytokine i nterleukin ( IL ) - 10 in the serum and a preferential appearance of regulatory type 2 macrophages in the patients’ bone marrow. Altogether, this data confirmed previous findings from in vitro and animal model studies regarding the MSCs ’ impact on myeloid cell populations . Driven by these ob servations we sought out to assess whether MSCs induce so - called myeloid derived suppressor cells (MDSCs) in aGVHD patients. Although we did not find a n MSC - associate d effect , we were the first to i dentify monocytic CD14 + HLA - DR low/neg MDSCs accumulating af ter allogeneic hematopoietic transplantation. We characterized their suppressive function ( via indoleamine - 2,3 - dioxygenase) and established a significant association with inflammatory cytokines and aGVHD. In fact, our data indicates that MDSCs are part of an immune regulating feedback mechanism that is activated during hyper - inflammations (such as in aGVHD). Overall, our results indicate that immune regulatory populations play a decisive role in various inflammatory diseases and MSCs could boost their respo nse s . Furthermore our work suggests that combining basic and translational research is pre - requisite for understanding the MSCs’ multifaceted interactions and for optimizing their clinical use

Early quantification of anti-CD19 CAR T cells by flow cytometry predicts response in R/R DLBCL

CAR T cells; Flow cytometryCèl·lules T CAR; Citometria de fluxCélulas T CAR; Citometría de flujoThe work was supported by a German Research Council research grant provided within the Sonderforschungsbereich Transregio SFB-TRR 388/1 2021 452881907, and German Research Council research grant 451580403 (M.S.). The work was further supported by the Bavarian Elite Graduate Training Network (M.S. and G.B.), the Wilhelm Sander-Stiftung (M.S.) (project no. 2018.087.1), the Else Kröner-Fresenius-Stiftung (V.B., K.R., V.L.B., and M.S.), the German Cancer Consortium (V.B.), the German Cancer Aid (F.M.) (grant 70113695), the Interdisciplinary Center for Clinical Research at the University Hospital of the University of Erlangen-Nuremberg (S.V.) (project no. D43), and the Bavarian Center for Cancer Research

Mesenchymal Stromal Cells Engage Complement and Complement Receptor Bearing Innate Effector Cells to Modulate Immune Responses

Infusion of human third-party mesenchymal stromal cells (MSCs) appears to be a promising therapy for acute graft-versus-host disease (aGvHD). To date, little is known about how MSCs interact with the body's innate immune system after clinical infusion. This study shows, that exposure of MSCs to blood type ABO-matched human blood activates the complement system, which triggers complement-mediated lymphoid and myeloid effector cell activation in blood. We found deposition of complement component C3-derived fragments iC3b and C3dg on MSCs and fluid-phase generation of the chemotactic anaphylatoxins C3a and C5a. MSCs bound low amounts of immunoglobulins and lacked expression of complement regulatory proteins MCP (CD46) and DAF (CD55), but were protected from complement lysis via expression of protectin (CD59). Cell-surface-opsonization and anaphylatoxin-formation triggered complement receptor 3 (CD11b/CD18)-mediated effector cell activation in blood. The complement-activating properties of individual MSCs were furthermore correlated with their potency to inhibit PBMC-proliferation in vitro, and both effector cell activation and the immunosuppressive effect could be blocked either by using complement inhibitor Compstatin or by depletion of CD14/CD11b-high myeloid effector cells from mixed lymphocyte reactions. Our study demonstrates for the first time a major role of the complement system in governing the immunomodulatory activity of MSCs and elucidates how complement activation mediates the interaction with other immune cells

Parvovirus-B19-Infektionen bei pädiatrischen Patienten mit hämatologischen und, oder onkologischen Erkrankungen

Parvovirus B19 (B19V)-Infektionen verlaufen bei Personen mit hämatologischen oder onkologischen Grunderkrankungen wesentlich schwerer als bei Gesunden. Um die Häufigkeit und die Auswirkungen einer B19V-Infektion bei pädiatrischen Patienten auf den Verlauf und die Therapie von hämatologischen und / oder onkologischen Erkrankungen zu bestimmen, wurden von 110 Kindern jeweils Serum und Knochenmarkproben auf Marker einer akuten, abgelaufenen oder persistierenden B19V-Infektion mittels qPCR, ELISA und WesternLine untersucht. 27/110 (24.5%) der Kinder litten an nicht malignen Erkrankungen (Anämie, Panzytopenie, Autoimmunerkrankungen). 68/110 (61.8%) Patienten waren an Leukämien, Lymphomen oder soliden, malignen Tumoren erkrankt. 15/110 Patienten (13.6%) wurden zunächst mit Verdacht auf eine maligne oder hämatologische Erkrankung aufgenommen, die jedoch im Verlauf ausgeschlossen wurde. Zum Zeitpunkt der Aufnahme wurden B19V-spezifisches IgM und IgG als Indikatoren einer akuten oder stattgefundenen B19V-Infektion bei fünf (4.5%) bzw. 48 Patienten (43.6%) festgestellt. B19V-DNA (103-109 geq/ml) wurde im Serum und / oder Knochenmark von 22 Patienten (20.0%) nachgewiesen. Davon litten fünf Kinder an Leukämien, zwei an einem Non-Hodgkin-Lymphom, sechs Kinder an soliden Tumoren, vier an Autoimmunerkrankungen und vier an hämatologischen Erkrankungen und ein Kind an Fieber unklarer Genese. Während der klinischen Beobachtung entwickelten vier weitere Leukämiepatienten eine Virämie. Eine persistierende B19V-Infektion wurde bei 13/22 DNA-positiven Patienten beobachtet. Die Therapie von B19V-DNA positiven Krebspatienten war mit einer erhöhten Supportivtherapie (Erythrozyten-, Thrombozytenkonzentraten und/oder Antibiotikatherapie) assoziiert. Eine akute B19V-Infektion wurde häufig bei Patienten mit hämatologischen und/oder onkologischen Erkrankungen beobachtet. Bei Patienten mit nicht malignen Erkrankungen wurden Anämien oder Autoimmunerkrankungen in Zusammenhang mit einer B19V-Infektion diagnostiziert. Weiterhin zeigte eine signifikante Anzahl von Krebspatienten Marker für eine akute, abgelaufene oder persistierende B19V-Infektion. Dies könnte durch die häufigen Therapien mit Blutprodukten in der Kombination mit einer therapeutischen Immunsuppression begünstigt werden. Bei B19V-infizierten Krebspatienten war eine komplexere Supportivtherapie erforderlich

CD33/CD3-bispecific T-cell engaging (BiTE®) antibody construct targets monocytic AML myeloid-derived suppressor cells

Abstract Acute myeloid leukemia (AML) is the most common acute leukemia amongst adults with a 5-year overall survival lower than 30%. Emerging evidence suggest that immune alterations favor leukemogenesis and/or AML relapse thereby negatively impacting disease outcome. Over the last years myeloid derived suppressor cells (MDSCs) have been gaining momentum in the field of cancer research. MDSCs are a heterogeneous cell population morphologically resembling either monocytes or granulocytes and sharing some key features including myeloid origin, aberrant (immature) phenotype, and immunosuppressive activity. Increasing evidence suggests that accumulating MDSCs are involved in hampering anti-tumor immune responses and immune-based therapies. Here, we demonstrate increased frequencies of CD14+ monocytic MDSCs in newly diagnosed AML that co-express CD33 but lack HLA-DR (HLA-DRlo). AML-blasts induce HLA-DRlo cells from healthy donor-derived monocytes in vitro that suppress T-cells and express indoleamine-2,3-dioxygenase (IDO). We investigated whether a CD33/CD3-bispecific BiTE® antibody construct (AMG 330) with pre-clinical activity against AML-blasts by redirection of T-cells can eradicate CD33+ MDSCs. In fact, T-cells eliminate IDO+CD33+ MDSCs in the presence of AMG 330. Depletion of total CD14+ cells (including MDSCs) in peripheral blood mononuclear cells from AML patients did not enhance AMG 330-triggered T-cell activation and expansion, but boosted AML-blast lysis. This finding was corroborated in experiments showing that adding MDSCs into co-cultures of T- and AML-cells reduced AML-blast killing, while IDO inhibition promotes AMG 330-mediated clearance of AML-blasts. Taken together, our results suggest that AMG 330 may achieve anti-leukemic efficacy not only through T-cell-mediated cytotoxicity against AML-blasts but also against CD33+ MDSCs, suggesting that it is worth exploring the predictive role of MDSCs for responsiveness towards an AMG 330-based therapy

CLL-cells induce IDOhi CD14+HLA-DRlo myeloid derived suppressor cells that inhibit T-cell responses and promote TRegs

Myeloid derived suppressor cells (MDSCs) represent a heterogeneous population that shares certain characteristics including an aberrant myeloid phenotype and the ability to suppress T-cells. MDSCs have been predominantly studied in malignant diseases and findings suggest involvement in tumor-associated immune suppression. Chronic lymphocytic leukemia (CLL) is the leukemia with the highest incidence among adults. Immune defects occur already at early disease stages and impact the clinical course. We assessed presence, frequency, association to other immune parameters, and functional properties of circulating CD14+ cells lacking HLA-DR expression (HLA-DRlo) in patients with untreated CLL. These monocytic cells represent one of the best-defined human MDSC subsets. Frequency of CD14+HLA-DRlo cells was significantly increased in CLL patients. Furthermore, MDSCs suppressed in vitro T-cell activation and induced suppressive regulatory T-cells (TRegs). The MDSC-mediated modulation of T-cells could be attributed to their increased indoleamine 2,3-dioxygenase (IDO) activity. CLL-cells induced IDOhi MDSCs from healthy donor monocytes suggesting bidirectional crosstalk between CLL-cells, MDSCs, and TRegs. Overall, we identified a MDSC population that expands in CLL. The exact mechanisms responsible for such accumulation remain to be elucidated and it will be of interest to test whether antagonizing suppressive functions of CLL MDSCs could represent a mean for enhancing immune responses

Contact-Dependent Depletion of Hydrogen Peroxide by Catalase Is a Novel Mechanism of Myeloid-Derived Suppressor Cell Induction Operating in Human Hepatic Stellate Cells

Abstract Myeloid-derived suppressor cells (MDSC) represent a unique cell population with distinct immunosuppressive properties that have been demonstrated to shape the outcome of malignant diseases. Recently, human hepatic stellate cells (HSC) have been reported to induce monocytic-MDSC from mature CD14+ monocytes in a contact-dependent manner. We now report a novel and unexpected mechanism by which CD14+HLADRlow/− suppressive cells are induced by catalase-mediated depletion of hydrogen peroxide (H2O2). Incubation of CD14+ monocytes with catalase led to a significant induction of functional MDSC compared with media alone, and H2O2 levels inversely correlated with MDSC frequency (r = −0.6555, p &amp;lt; 0.05). Catalase was detected in primary HSC and a stromal cell line, and addition of the competitive catalase inhibitor hydroxylamine resulted in a dose-dependent impairment of MDSC induction and concomitant increase of H2O2 levels. The NADPH-oxidase subunit gp91 was significantly increased in catalase-induced MDSC as determined by quantitative PCR outlining the importance of oxidative burst for the induction of MDSC. These findings represent a so far unrecognized link between immunosuppression by MDSC and metabolism. Moreover, this mechanism potentially explains how stromal cells can induce a favorable immunological microenvironment in the context of tissue oxidative stress such as occurs during cancer therapy.</jats:p

Accumulation of T-cell-suppressive PD-L1high extracellular vesicles is associated with GvHD and might impact GvL efficacy

Allogeneic hematopoietic stem cell transplantation (allo-HSCT) represents the only curative treatment option for a number of hemato-oncological disorders. In fact, allo-HSCT is considered as one of the most successful immunotherapies as its clinical efficacy is based on the donor T-cells’ capacity to control residual disease. This process is known as the graft-versus-leukemia (GvL) reaction. However, alloreactive T-cells can also recognize the host as foreign and trigger a systemic potentially life-threatening inflammatory disorder termed graft-versus-host disease (GvHD). A better understanding of the underlying mechanisms that lead to GvHD or disease relapse could help us to improve efficacy and safety of allo-HSCT. In recent years, extracellular vesicles (EVs) have emerged as critical components of intercellular crosstalk. Cancer-associated EVs that express the immune checkpoint molecule programmed death-ligand 1 (PD-L1) can suppress T-cell responses and thus contribute to immune escape. At the same time, it has been observed that inflammation triggers PD-L1 expression as part of a negative feedback network.Here, we investigated whether circulating EVs following allo-HSCT express PD-L1 and tested their efficacy to suppress the ability of (autologous) T-cells to effectively target AML blasts. Finally, we assessed the link between PD-L1 levels on EVs to (T-)cell reconstitution, GvHD, and disease relapse.We were able to detect PD-L1+ EVs that reached a peak PD-L1 expression at 6 weeks post allo-HSCT. Development of acute GvHD was linked to the emergence of PD-L1high EVs following allo-HSCT. Moreover, PD-L1 levels correlated positively with GvHD grade and declined (only) on successful therapeutic intervention. T-cell-inhibitory capacity was higher in PD-L1high EVs as compared with their PD-L1low counterparts and could be antagonized using PD-L1/PD-1 blocking antibodies. Abundance of T-cell-suppressive PD-L1high EVs appears to also impact GvL efficacy as patients were at higher risk for relapse. Finally, patients of PD-L1high cohort displayed a reduced overall survival.Taken together, we show that PD-L1-expressing EVs are present following allo-HSCT. PD-L1 levels on EVs correlate with their ability to suppress T-cells and the occurrence of GvHD. The latter observation may indicate a negative feedback mechanism to control inflammatory (GvHD) activity. This intrinsic immunosuppression could subsequently promote disease relapse