Methotrexate enhances the anti-inflammatory effect of CF101 via up-regulation of the A3 adenosine receptor expression.
Methotrexate (MTX) exerts an anti-inflammatory effect via its metabolite adenosine, which activates adenosine receptors. The A3 adenosine receptor (A3AR) was found to be highly expressed in inflammatory tissues and peripheral blood mononuclear cells (PBMCs) of rats with adjuvant-induced arthritis (AIA). CF101 (IB-MECA), an A3AR agonist, was previously found to inhibit the clinical and pathological manifestations of AIA. The aim of the present study was to examine the effect of MTX on A3AR expression level and the efficacy of combined treatment with CF101 and MTX in AIA rats. AIA rats were treated with MTX, CF101, or both agents combined. A3AR mRNA, protein expression and exhibition were tested in paw and PBMC extracts from AIA rats utilizing immunohistochemistry staining, RT-PCR and Western blot analysis. A3AR level was tested in PBMC extracts from patients chronically treated with MTX and healthy individuals. The effect of CF101, MTX and combined treatment on A3AR expression level was also tested in PHA-stimulated PBMCs from healthy individuals and from MTX-treated patients with rheumatoid arthritis (RA). Combined treatment with CF101 and MTX resulted in an additive anti-inflammatory effect in AIA rats. MTX induced A2AAR and A3AR over-expression in paw cells from treated animals. Moreover, increased A3AR expression level was detected in PBMCs from MTX-treated RA patients compared with cells from healthy individuals. MTX also increased the protein expression level of PHA-stimulated PBMCs from healthy individuals. The increase in A3AR level was counteracted in vitro by adenosine deaminase and mimicked in vivo by dipyridamole, demonstrating that receptor over-expression was mediated by adenosine. In conclusion, the data presented here indicate that MTX induces increased A3AR expression and exhibition, thereby potentiating the inhibitory effect of CF101 and supporting combined use of these drugs to treat RA.
Ochaion
Avivit
A
Can-Fite Biopharma Ltd, 10 Bareket Street, Kiryat-Matalon, Petah-Tikva, 49170, Israel.
Bar-Yehuda
Sara
S
Cohn
Shira
S
Del Valle
Luis
L
Perez-Liz
Georginia
G
Madi
Lea
L
Barer
Faina
F
Farbstein
Motti
M
Fishman-Furman
Sari
S
Reitblat
Tatiana
T
Reitblat
Alexander
A
Amital
Howard
H
Levi
Yair
Y
Molad
Yair
Y
Mader
Reuven
R
Tishler
Moshe
M
Langevitz
Pnina
P
Zabutti
Alexander
A
Fishman
Pnina
P
D000241:Adenosine / Q000031:analogs & derivatives* / Q000494:pharmacology; D000818:Animals; D000893:Anti-Inflammatory Agents / Q000494:pharmacology*; D001168:Arthritis / Q000188:drug therapy* / Q000378:metabolism* / Q000473:pathology; D015153:Blotting, Western; D004359:Drug Therapy, Combination; D005260:Female; D006801:Humans; D007150:Immunohistochemistry; D007963:Leukocytes, Mononuclear / Q000187:drug effects / Q000378:metabolism; D008727:Methotrexate / Q000494:pharmacology*; D008875:Middle Aged; D051381:Rats; D011917:Rats, Inbred Lew; D043684:Receptor, Adenosine A3 / Q000096:biosynthesis / Q000187:drug effects*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015854:Up-Regulation
D000893:Anti-Inflammatory Agents; C478920:CF101; D043684:Receptor, Adenosine A3; D000241:Adenosine; D008727:Methotrexate