Quantification of T‐cell–mediated apoptosis in heterogeneous leukemia populations using four‐color multiparameter flow cytometry

Westers, Theresia M.; Houtenbos, Ilse; Schuurhuis, Gerrit Jan; Ossenkoppele, Gert J.; Loosdrecht, Arjan A. van de

doi:10.1002/cyto.a.20146

Cited by 15 publications

(15 citation statements)

References 23 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Although not yet proven in clinical studies, in vitro data that we and others have generated show that AML-DC possess all the prerequisite functions needed to elicit an immune response in vivo (14,16,17). In a phase I pilot study on chronic myelogenous leukemia -derived dendritic cell vaccination in the advanced-stage disease, delayed type hypersensitivity responses representing autologous chronic myelogenous leukemia -specific T-cell responses were detected (27) However, clinical dendritic cell vaccination studies have, until now, shown limited success (18,19).…”

Section: Discussionmentioning

confidence: 91%

See 1 more Smart Citation

Leukemia-Specific T-Cell Reactivity Induced by Leukemic Dendritic Cells Is Augmented by 4-1BB Targeting

Houtenbos¹,

Westers²,

Dijkhuis³

et al. 2007

Clinical Cancer Research

Self Cite

View full text Add to dashboard Cite

Purpose: Acute myelogenous leukemia (AML) blasts are able to differentiate into leukemiaderived dendritic cells (AML-DC), thereby enabling efficient presentation of known and unknown leukemic antigens. Advances in culture techniques and AML-DC characterization justify clinical application. However, additional measures are likely needed to potentiate vaccines and overcome the intrinsic tolerant state of the patients' immune system. Engagement of the costimulatory molecule 4-1BB can break immunologic tolerance and increase CTL responses. In this study, we examined the role of the 4-1BB ligand (4-1BBL) onT-cell responses induced by AML-DC. Experimental Design: In allogeneic and autologous cocultures of T cells and AML-DC, the effect of the addition of 4-1BBL on T-cell proliferation, T-cell subpopulations, and T-cell function was determined. Results: Addition of 4-1BBL to cocultures of AML-DC and Tcells induced a preferential increase in the proliferation of CD8 + T cells. Increased differentiation into effector and central memory populations was observed in both CD4 + and CD8 + T cells in the presence of 4-1BBL. AML-DC induce a T helper 1 response, characterized by high IFN-g production, which is significantly increased by targeting 4-1BB. T cells primed in the presence of 4-1BBL show specificity for the leukemia-associated antigen Wilms' tumor 1, whereas cytotoxicity assays with leukemic blast targets showed the cytolytic potential of Tcells primed in the presence of 4-1BBL. Conclusion: We conclude that 4-1BBL is an effective adjuvant to enhance T-cell responses elicited by AML-DC.

show abstract

Section: Discussionmentioning

confidence: 91%

“…The ability of cultured T cells to kill leukemic blasts was evaluated in a flow cytometric cytotoxicity assay, as previously described (17). Briefly, AML-DC -stimulated T cells (effector cells) with or without 4-1BBL were cultured with corresponding AML blasts (target cells) at different effector to target cell ratios.…”

Section: Methodsmentioning

confidence: 99%

Leukemia-Specific T-Cell Reactivity Induced by Leukemic Dendritic Cells Is Augmented by 4-1BB Targeting

Houtenbos¹,

Westers²,

Dijkhuis³

et al. 2007

Clinical Cancer Research

Self Cite

View full text Add to dashboard Cite

show abstract

“…Leukemic cells were harvested and taken up in PBS and apoptosis was induced by heat shock (2 h at 42°C) or by incubation for 2 h at 37°C with ARA-C (10 μg/ml; Mayne Pharma, Warwickshire, UK). The percentage of necrotic, apoptotic, and viable cells was determined before and after apoptosis induction by incubation with Syto-62 (5 nM; Molecular Probes, Eugene, OR), PSC833 (2 μM; Novartis, Basel, Switzerland) and 7-aminoactinomycin D (7-AAD; ViaProbe, Pharmingen, San Diego, CA) for 45 min at 37°C, and by flow cytometric analysis as described previously [28]. Syto-62 is a fluorescent nucleic acid stain that exhibits bright fluorescence upon binding to nucleic acids and is retained in viable cells.…”

Section: Methodsmentioning

confidence: 99%

Targeting Toll-like receptor 7/8 enhances uptake of apoptotic leukemic cells by monocyte-derived dendritic cells but interferes with subsequent cytokine-induced maturation

Ancker

Luijn

Ruben

et al. 2010

Cancer Immunol Immunother

Self Cite

View full text Add to dashboard Cite

Therapeutic vaccination with dendritic cells (DC) is an emerging investigational therapy for eradication of minimal residual disease in acute myeloid leukemia. Various strategies are being explored in manufacturing DC vaccines ex vivo, e.g., monocyte-derived DC (MoDC) loaded with leukemia-associated antigens (LAA). However, the optimal source of LAA and the choice of DC-activating stimuli are still not well defined. Here, loading with leukemic cell preparations (harboring both unknown and known LAA) was explored in combination with a DC maturation-inducing cytokine cocktail (CC; IL-1β, IL-6, TNF-α, and PGE2) and Toll-like receptor ligands (TLR-L) to optimize uptake. Since heat shock induced apoptotic blasts were more efficiently taken up than lysates, we focused on uptake of apoptotic leukemic cells. Uptake of apoptotic blast was further enhanced by the TLR7/8-L R848 (20–30%); in contrast, CC-induced maturation inhibited uptake. CC, and to a lesser extent R848, enhanced the ability of MoDC to migrate and stimulate T cells. Furthermore, class II-associated invariant chain peptide expression was down-modulated after R848- or CC-induced maturation, indicating enhanced processing and presentation of antigenic peptides. To improve both uptake and maturation, leukemic cells and MoDC were co-incubated with R848 for 24 h followed by addition of CC. However, this approach interfered with CC-mediated MoDC maturation as indicated by diminished migratory and T cell stimulatory capacity, and the absence of IL-12 production. Taken together, our data demonstrate that even though R848 improved uptake of apoptotic leukemic cells, the sequential use of R848 and CC is counter-indicated due to its adverse effects on MoDC maturation.

show abstract

“…22 Peripheral CD8 + T cells (effector cells) and bone marrow derived CD34 + blasts (target cells) were selected by magnetic separation (positive selection with CD8 and CD34 microbeads respectively, Miltenyi Biotec, Bergisch Gladbach, Germany). During this procedure also CD8 dim cells were selected resulting in effector population consisting of CD3 +…”

Section: Flow Cytometric Cytotoxic Assaymentioning

confidence: 99%

Immune mediated autologous cytotoxicity against hematopoietic precursor cells in patients with myelodysplastic syndrome

Chamuleau¹,

Westers²,

Dreunen³

et al. 2009

Haematologica

View full text Add to dashboard Cite

Quantification of T‐cell–mediated apoptosis in heterogeneous leukemia populations using four‐color multiparameter flow cytometry

Cited by 15 publications

References 23 publications

Leukemia-Specific T-Cell Reactivity Induced by Leukemic Dendritic Cells Is Augmented by 4-1BB Targeting

Leukemia-Specific T-Cell Reactivity Induced by Leukemic Dendritic Cells Is Augmented by 4-1BB Targeting

Targeting Toll-like receptor 7/8 enhances uptake of apoptotic leukemic cells by monocyte-derived dendritic cells but interferes with subsequent cytokine-induced maturation

Immune mediated autologous cytotoxicity against hematopoietic precursor cells in patients with myelodysplastic syndrome

Contact Info

Product

Resources

About