In vitro evaluation of adenosine 5′-monophosphate as an imaging agent of tumor metabolism

Adenosine appears to play an important role in tumor growth and metastasis. Synthesized ¹¹C-adenosine 5′-monophosphate (AMP) has recently been reported as a potential tumor-imaging radiotracer. Methods: A variety of human tumor cell lines (SKOV-3, SCC-15, U251, U87, Raji, and Daudi) were incubated with 3.7 kBq (0.1 μCi) of [2-³H]AMP (³H-AMP), [5,6-³H]FDG (³H-FDG), or [2-³H]adenosine (³H-adenosine) in low-physiologicglucose serum-free medium. Selected cells were exposed to caffeine, dipyridamole, adenosine 5′-(α, β-methylene)diphosphate (APCP), or unlabeled adenosine before exposure to the radiotracer. R-phycoerythrin-conjugated mouse antihuman monoclonal antibody to human CD73 was used for immunophenotyping. High-performance liquid chromatography was used to characterize the intracellular metabolites of ³H-AMP after intracellular uptake. Results: Intracellular uptake of ³H-AMP was significant - 10 to 100 times the uptake of ³H-FDG, depending on the particular tumor cell line. Preexposure of SKOV-3 cells to caffeine, a competitive inhibitor of adenosine receptors, did not affect cellular uptake ofv ³H-AMP. However, preexposure of SKOV-3 cells to dipyridamole, an equilibrative nucleoside transporter inhibitor; APCP, a CD73 (ecto-5′-nucleotidase) inhibitor; or cold adenosine significantly inhibited cellular uptake of ³H-AMP. SKOV-3 uptake of ³H-adenosine was inhibited by dipyridamole but not APCP. U251 uptake of ³H-AMP was significantly inhibited by dipyridamole and APCP. U87 uptake of ³H-AMP was only partially inhibited by dipyridamole and APCP. However, Raji and Daudi cells had significantly lower uptake of ³H-AMP than of ³H-FDG but had significantly increased uptake of ³H-adenosine, whichwas inhibited by dipyridamole.Raji and Daudi cells were negative, but the SKOV-3 cells positive, for CD73 cell-surface expression. ³H-Adenosine metabolites were persistently retained after influx into the cell, predominantly as ³H-adenosine triphosphate and ³H-adenosine diphosphate. Conclusion: Cancer cell lines evaluated in vitro had significantly elevated uptake of radiolabeled AMP, on the order of 10- to100-fold, in comparison to radiolabeled FDG. The mechanism of intracellular uptake depends predominantly on equilibrative nucleoside transporters after conversion of AMP to adenosine by CD73 in SKOV-3, SCC-15, and U251 cells. Intracellular retention is due to phosphorylation to adenosine triphosphate and adenosine diphosphate. Raji and Daudi cells have low uptake of radiolabeled AMP because of a lack of CD73 expression. This in vitro evaluation using ³H-AMP with tumor cell lines supports thepotential of ¹¹C-AMP for use in targeting the nucleoside transport pathway in PET imaging of tumors.