ABSTRACT
Background: In rheumatoid arthritis, articular inflammation is a hallmark of disease, while the involvement of extra-articular tissues is less well defined. Here, we examined the feasibility of PET imaging with the macrophage tracer [18F]fluoro-PEG-folate, targeting folate receptor ß (FRß), to monitor systemic inflammatory disease in liver and spleen of arthritic rats before and after methotrexate (MTX) treatment. Methods: [18F]Fluoro-PEG-folate PET scans (60 min) were acquired in saline- and MTX-treated (1 mg/kg, 4x) arthritic rats, followed by tissue resection and radiotracer distribution analysis. Liver and spleen tissues were stained for ED1/ED2-macrophage markers and FRß expression. Results: [18F]Fluoro-PEG-folate PET and ex vivo tissue distribution studies revealed a significant (p < 0.01) 2-fold lower tracer uptake in both liver and spleen of MTX-treated arthritic rats. Consistently, ED1- and ED2-positive macrophages were significantly (p < 0.01) decreased in liver (4-fold) and spleen (3-fold) of MTX-treated compared with saline-treated rats. Additionally, FRß-positive macrophages were also significantly reduced in liver (5-fold, p < 0.005) and spleen (3-fold, p < 0.01) of MTX- versus saline-treated rats. Conclusions: MTX treatment reduced activated macrophages in liver and spleen, as markers for systemic inflammation in these organs. Macrophage PET imaging with [18F]fluoro-PEG-folate holds promise for detection of systemic inflammation in RA as well as therapy (MTX) response monitoring.
Subject(s)
Arthritis, Experimental , Arthritis, Rheumatoid , Fluorine Radioisotopes/pharmacology , Folic Acid/analogs & derivatives , Methotrexate/pharmacology , Polyethylene Glycols/pharmacology , Positron-Emission Tomography , Animals , Arthritis, Experimental/chemically induced , Arthritis, Experimental/diagnostic imaging , Arthritis, Experimental/metabolism , Arthritis, Rheumatoid/chemically induced , Arthritis, Rheumatoid/diagnostic imaging , Arthritis, Rheumatoid/metabolism , Folic Acid/pharmacology , Inflammation/chemically induced , Inflammation/diagnostic imaging , Inflammation/metabolism , Macrophages/metabolism , Macrophages/pathology , Methotrexate/pharmacokinetics , Rats , Rats, WistarABSTRACT
Aminopeptidase inhibitors are receiving attention as combination chemotherapeutic agents for the treatment of refractory acute myeloid leukemia. However, the factors determining therapeutic efficacy remain elusive. Here we identified the molecular basis of acquired resistance to CHR2863, an orally available hydrophobic aminopeptidase inhibitor prodrug with an esterase-sensitive motif, in myeloid leukemia cells. CHR2863 enters cells by diffusion and is retained therein upon esterase activity-mediated conversion to its hydrophilic active metabolite drug CHR6768, thereby exerting amino acid depletion. Carboxylesterases (CES) serve as candidate prodrug activating enzymes given CES1 expression in acute myeloid leukemia specimens. We established two novel myeloid leukemia sublines U937/CHR2863(200) and U937/CHR2863(5uM), with low (14-fold) and high level (270-fold) CHR2863 resistance. The latter drug resistant cells displayed: (i) complete loss of CES1-mediated drug activation associated with down-regulation of CES1 mRNA and protein, (ii) marked retention/sequestration of the prodrug, (iii) a substantial increase in intracellular lipid droplets, and (iv) a dominant activation of the pro-survival Akt/mTOR pathway. Remarkably, the latter feature coincided with a gain of sensitivity to the mTOR inhibitor rapamycin. These finding delineate the molecular basis of CHR2863 resistance and offer a novel modality to overcome this drug resistance in myeloid leukemia cells.
