An Optimized CFSE-based T-cell Suppression Assay to Evaluate the Suppressive Capacity of Regulatory T-Cells Induced by Human Tolerogenic Dendritic Cells

Abstract Abstract 2156 Multiple approaches for treatment of malignant disease presently aim to combine targeted therapy with tyrosine kinase inhibitors (TKI) with immunotherapy. Dendritic cells (DC) are frequently used in such strategies due to their unique ability to initiate potent T cell anti-tumor immunity. Unfortunately, DC may also activate suppressive CD25+FOXP3+ regulatory T cells (Treg), which depends on the stimuli that influence DC in immature state and/or during development from precursor cells. High frequencies of Treg have been described in several types of tumors within the tumor microenvironment, which is associated with poor prognosis and reduced survival. DC development and function are moreover governed by various tyrosine kinases of which some are also inhibited by clinically used TKI. TKI thus may cause immunoinhibitory side effects, and we previously demonstrated that exposure of monocyte-derived DC to the BCR-ABL inhibitor imatinib causes up-regulation of the immunosuppressive type I transmembrane glycoprotein osteoactivin (GPNMB, DC-HIL) and reduces expression of activating surface antigens as well as T cell-stimulatory capacity of DC in vitro (Schwarzbich et al., 2012). Other investigators reported that imatinib induces functionally Treg in CML patients, but the underlying mechanisms are so far unknown. (Bachy et al., 2011). On the other hand, TKI may inhibit proliferation and suppressive capacity of regulatory T cells in vitro (Chen et al., 2007). Here we tried to solve this apparent discrepancy by analyzing the influence of TKI on DC-Treg interaction. Monocyte-derived DC (moDC) were generated over 7 days by exposing blood monocytes to GM-CSF and IL-4. TNF was added on day 6 of culture in case of maturation, and imatinib or nilotinib (3μM each) were added to the culture medium every second day starting from the first day of culture. Induction and functionality of Treg was determined by FACS and so called effector T cell suppression assays upon culture of moDC with autologous PBMC. We found that exposure of moDC to imatinib or nilotinib only slightly increased the frequency of Treg as compared to controls. However, these Treg strongly inhibited autologous T cell proliferation as assessed by T cell suppression assays. This was mediated by direct cellular interaction, as culture supernatants of TKI-treated DC did not alter Treg function and also did not contain elevated levels of the immunosuppressive (and Treg inducing) cytokines TGF-β and IL-10. Thus, our data indicate that the seemingly contradictory results of the in vivo and in vitro studies described above may be explained by the effects caused by exposure of moDC to BCR-ABL TKI which results in the induction of functionally active Treg. These findings are of special importance for future combinatory approaches using TKI and DC-based immunotherapy. Disclosures: No relevant conflicts of interest to declare.

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Both Maturation and Survival of Human Dendritic Cells are Impaired in the Presence of Anergic/Suppressor T Cells

Clinical and Developmental Immunology ◽

10.1080/10446670310001593505 ◽

2003 ◽

Vol 10 (1) ◽

pp. 61-65 ◽

Cited By ~ 1

Author(s):

L. Frasca ◽

C. Scottà ◽

G. Lombardi ◽

E. Piccolella

Keyword(s):

T Cells ◽

Dendritic Cells ◽

T Cell ◽

T Cell Activation ◽

Cell Activation ◽

Costimulatory Molecule ◽

T Cell Suppression ◽

Cell Suppression ◽

Anergic T Cells ◽

Human Dendritic Cells

T cell suppression is a well established phenomenon, but the mechanisms involved are still a matter of debate. Mouse anergic T cells were shown to suppress responder T cell activation by inhibiting the antigen presenting function of DC. In the present work we studied the effects of co-culturing human anergic CD4+T cells with autologous dendritic cells (DC) at different stages of maturation. Either DC maturation or survival, depending on whether immature or mature DC where used as APC, was impaired in the presence of anergic cells. Indeed, MHC and costimulatory molecule up-regulation was inhibited in immature DC, whereas apoptotic phenomena were favored in mature DC and consequently in responder T cells. Defective ligation of CD40 by CD40L (CD154) was responsible for CD95-mediated and spontaneous apoptosis of DC as well as for a failure of their maturation process. These findings indicate that lack of activation of CD40 on DC by CD40L-defective anergic cells might be the primary event involved in T cell suppression and support the role of CD40 signaling in regulating both activation and survival of DC.

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CD4+Foxp3+ regulatory T cells mediate Toxoplasma gondii-induced T-cell suppression through an IL-2-related mechanism but independently of IL-10

European Journal of Immunology ◽

10.1002/eji.201141507 ◽

2011 ◽

Vol 41 (12) ◽

pp. 3529-3541 ◽

Cited By ~ 19

Author(s):

Eda P. Tenorio ◽

Jacquelina Fernández ◽

Carlos Castellanos ◽

Jonadab E. Olguín ◽

Rafael Saavedra

Keyword(s):

T Cells ◽

T Cell ◽

Toxoplasma Gondii ◽

Regulatory T Cells ◽

T Cell Suppression ◽

Cell Suppression

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Abatacept (CTLA-4Ig) therapy prevents activation induced cell death (AICD) of regulatory T cells and reduces the susceptibility of T cells to regulatory T cell suppression in patients with rheumatoid arthritis (RA)

Annals of the Rheumatic Diseases ◽

10.1136/annrheumdis-2011-201230.22 ◽

2012 ◽

Vol 71 (Suppl 1) ◽

pp. A10.1-A10 ◽

Cited By ~ 1

Author(s):

M Bonelli ◽

L Göschl ◽

S Blüml ◽

J S Smolen ◽

C Scheinecker

Keyword(s):

Rheumatoid Arthritis ◽

T Cells ◽

Cell Death ◽

T Cell ◽

Regulatory T Cells ◽

Regulatory T Cell ◽

Activation Induced Cell Death ◽

T Cell Suppression ◽

Cell Suppression

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Expansion of regulatory T cells and increased expression of programmed death-1 contribute to liver T cell suppression due to obstructive jaundice

Journal of the American College of Surgeons ◽

10.1016/j.jamcollsurg.2010.06.022 ◽

2010 ◽

Vol 211 (3) ◽

pp. S11

Author(s):

Kristin M. Ryan ◽

Doreen Osgood ◽

Ponnandai Somasundar ◽

N. Joseph Espat ◽

Steven C. Katz ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Regulatory T Cells ◽

Obstructive Jaundice ◽

T Cell Suppression ◽

Programmed Death ◽

Programmed Death 1 ◽

Cell Suppression

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Antigen-specific T cell suppression by human CD4+CD25+ regulatory T cells

European Journal of Immunology ◽

10.1002/1521-4141(200206)32:6<1621::aid-immu1621>3.0.co;2-q ◽

2002 ◽

Vol 32 (6) ◽

pp. 1621 ◽

Cited By ~ 186

Author(s):

Leonie S. Taams ◽

Milica Vukmanovic-Stejic ◽

Jay Smith ◽

Padraic J. Dunne ◽

Jean M. Fletcher ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Regulatory T Cells ◽

T Cell Suppression ◽

Cell Suppression

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Humoral immune response to adenovirus induce tolerogenic bystander dendritic cells that promote generation of regulatory T cells

10.1101/334508 ◽

2018 ◽

Author(s):

Thi Thu Phuong Tran ◽

Karsten Eichholz ◽

Patrizia Amelio ◽

Crystal Moyer ◽

Glen R Nemerow ◽

...

Keyword(s):

T Cells ◽

Dendritic Cells ◽

T Cell ◽

Regulatory T Cells ◽

Immune Responses ◽

Persistent Infections ◽

Cell Immunity ◽

Tolerogenic Dendritic Cells ◽

Antigen Presenting ◽

Tolerogenic Dcs

AbstractFollowing repeated encounters with adenoviruses most of us develop robust humoral and cellular immune responses that are thought to act together to combat ongoing and subsequent infections. Yet in spite of robust immune responses, adenoviruses establish subclinical persistent infections that can last for decades. While adenovirus persistence pose minimal risk in B-cell compromised individuals, if T-cell immunity is severely compromised, reactivation of latent adenoviruses can be life threatening. This dichotomy led us to ask how anti-adenovirus antibodies influence adenovirus-specific T-cell immunity. Using primary human blood cells, transcriptome and secretome profiling, and pharmacological, biochemical, genetic, molecular, and cell biological approaches, we initially found that healthy adults harbor adenovirus-specific regulatory T cells (Tregs). As peripherally induced Tregsare generated by tolerogenic dendritic cells (DCs), we then addressed how tolerogenic DCs could be created. Here, we demonstrate that DCs that take up immunoglobulin-complexed (IC)-adenoviruses create an environment that causes bystander DCs to become tolerogenic. These adenovirus antigen-loaded tolerogenic DCs can drive naïve T cells to mature into adenovirus-specific Tregs. Our results may provide ways to improve antiviral therapy and/or pre-screening high-risk individuals undergoing immunosuppression.Author summaryWhile numerous studies have addressed the cellular and humoral response to primary virus encounters, relatively little is known about the interplay between persistent infections, neutralizing antibodies, antigen-presenting cells, and the T-cell response. Our studies suggests that if adenovirus–antibody complexes are taken up by professional antigen-presenting cells (dendritic cells), the DCs generate an environment that causes bystander dendritic cells to become tolerogenic. These tolerogenic dendritic cells favors the creation of adenovirus-specific regulatory T cells. While this pathway likely favors pathogen survival, there may be advantages for the host also.

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