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Antiproliferative effects of selective adenosine receptor agonists and antagonists on human lymphocytes: evidence for receptor-independent mechanisms

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Abstract

The effects of standard adenosine receptor (AR) agonists and antagonists on the proliferation of human T lymphocytes, unstimulated and phytohemagglutinin-stimulated human peripheral blood lymphocytes (PBL), and Jurkat T cells were investigated. Real-time PCR measurements confirmed the presence of all four AR subtypes on the investigated cells, although at different expression levels. A2A ARs were predominantly expressed in PBL and further upregulated upon stimulation, while malignant Jurkat T cells showed high expression levels of A1, A2A, and A2B ARs. Cell proliferation was measured by [3H]-thymidine incorporation assays. Several ligands, including the subtype-selective agonists CPA (A1), BAY60-6583 (A2B), and IB-MECA (A3), and the antagonists PSB-36 (A1), MSX-2 (A2A), and PSB-10 (A3) significantly inhibited cell proliferation at micromolar concentrations, which were about three orders of magnitude higher than their AR affinities. In contrast, further investigated AR ligands, including the agonists NECA (nonselective) and CGS21680 (A2A), and the antagonists preladenant (SCH-420814, A2A), PSB-1115 (A2B), and PSB-603 (A2B) showed no or only minor effects on lymphocyte proliferation. The anti-proliferative effects of the AR agonists could not be blocked by the corresponding antagonists. The non-selective AR antagonist caffeine stimulated phytohemagglutinin-activated PBL with an EC50 value of 104 μM. This is the first study to compare a complete set of commonly used AR ligands for all subtypes on lymphocyte proliferation. Our results strongly suggest that these compounds induce an inhibition of lymphocyte proliferation and cell death through AR-independent mechanisms.

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Abbreviations

ADA:

Adenosine deaminase

AR:

Adenosine receptor

BAY60-6583:

2-[6-Amino-3,5-dicyano-4-[4-(cyclopropylmethoxy)phenyl]pyridin-2-ylsulfanyl]acetamide

cAMP:

Cyclic AMP

CGS-21680:

(2-p-[2-Carboxyethyl]phenethylamino)-5′-N-ethylcarboxamido-adenosine

CI-IB-MECA:

2-Chloro-N 6-(3-iodobenzyl)-9-[5-(methyl-carbamoyl)-β-d-ribofuranosyl]adenine

CPA:

N 6-Cyclopentyladenosine

DMSO:

Dimethyl sulfoxide

EDTA:

Ethylenediaminetetraacetic acid

FCS:

Fetal calf serum

GPCR(s):

G protein-coupled receptor(s)

IB-MECA:

N 6-(3-Iodobenzyl)-5′-N-methylcarboxamidoadenosine

MSX-2:

3-(3-Hydroxypropyl)-7-methyl-8-(m-methoxystyryl)-1-propargylxanthine

NECA:

5′-N-Ethylcarboxamidoadenosine

PBL:

Peripheral blood lymphocytes

PBS:

Phosphate-buffered saline

PCR:

Polymerase chain reaction

PHA:

Phytohemagglutinin

PSB-10:

(R)-8-Ethyl-4-methyl-2-(2,3,5-trichlorophenyl)-4,5,7,8-tetrahydro-1H-imidazo[2,1-i]purin-5-one

PSB-36:

1-Butyl-8-(3-noradamantanyl)-3-(3-hydroxypropyl)xanthine

PSB-603:

8-[4-[4-(4-Chlorophenyl)piperazine-1-sulfonyl)phenyl]]-1-propylxanthine

PSB-1115:

1-Propyl-8-p-sulfophenylxanthine

SCID:

Severe combined immunodeficiency

SCH420814:

Preladenant

TNF:

Tumor necrosis factor

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Acknowledgments

This study was supported by the Deutsche Forschungsgemeinschaft (DFG) within the Graduiertenkolleg GRK 677 (S.K.L, C.E.M.) and the NRW-Forschungsschule Biotech Pharma (A.C.S., C.E.M.). We thank H. Eltzschig and S. Zug (University of Tübingen, Germany) and A. Bill (University of Bonn, Germany) for their support in real-time PCR experiments. The technical assistance of N. Florin is gratefully acknowledged.

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Authors and Affiliations

  1. PharmaCenter Bonn, Pharmaceutical Institute, Pharmaceutical Sciences Bonn (PSB), Pharmaceutical Chemistry I, University of Bonn, An der Immenburg 4, 53121, Bonn, Germany

    Anke C. Schiedel, Svenja K. Lacher & Christa E. Müller

  2. Institute of Molecular Medicine and Experimental Immunology, Faculty of Medicine, University of Bonn, Bonn, Germany

    Carsten Linnemann & Percy A. Knolle

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  1. Anke C. Schiedel
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  2. Svenja K. Lacher
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Correspondence to Christa E. Müller.

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Anke C. Schiedel and Svenja K. Lacher contributed equally.

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Schiedel, A.C., Lacher, S.K., Linnemann, C. et al. Antiproliferative effects of selective adenosine receptor agonists and antagonists on human lymphocytes: evidence for receptor-independent mechanisms. Purinergic Signalling 9, 351–365 (2013). https://doi.org/10.1007/s11302-013-9354-7

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