Uveal melanoma cell growth is inhibited by aminoimidazole carboxamide ribonucleotide (AICAR) partially through activation of AMP-dependent kinase.
OBJECTIVE
To evaluate the effects and mechanism of aminoimidazole carboxamide ribonucleotide (AICAR), an AMP-dependent kinase (AMPK) activator, on the growth of uveal melanoma cell lines.


METHODS
Four different cell lines were treated with AICAR (1-4 mM). Cell growth was assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium (MTT) assay. Cell cycle analysis was conducted by flow cytometry; additionally, expression of cell-cycle control proteins, cell growth transcription factors, and downstream effectors of AMPK were determined by RT-PCR and Western blot.


RESULTS
Aminoimidazole carboxamide ribonucleotide inhibited cell growth, induced S-phase arrest, and led to AMPK activation. Aminoimidazole carboxamide ribonucleotide treatment was associated with inhibition of eukaryotic translation initiation factor 4E-BP1 phosphorylation, a marker of mammalian target of rapamycin (mTOR) pathway activity. Aminoimidazole carboxamide ribonucleotide treatment was also associated with downregulation of cyclins A and D, but had minimal effects on the phosphorylation of ribosomal protein S6 or levels of the macroautophagy marker LC3B. The effects of AICAR were abolished by treatment with dipyridamole, an adenosine transporter inhibitor that blocks the entry of AICAR into cells. Treatment with adenosine kinase inhibitor 5-iodotubericidin, which inhibits the conversion of AICAR to its 5'-phosphorylated ribotide 5-aminoimidazole-4-carboxamide-1-D-ribofuranosyl-5'-monophosphate (ZMP; the direct activator of AMPK), reversed most of the growth-inhibitory effects, indicating that some of AICAR's antiproliferative effects are mediated at least partially through AMPK activation.


CONCLUSIONS
Aminoimidazole carboxamide ribonucleotide inhibited uveal melanoma cell proliferation partially through activation of the AMPK pathway and downregulation of cyclins A1 and D1.
Retina Service, Angiogenesis Laboratory, Massachusetts Eye and Ear Infirmary, Department of Ophthalmology, Harvard Medical School, Boston, Massachusetts, United States.
Ahmad
A
Al-Moujahed
Pediatric Surgery Laboratories, Massachusetts General Hospital, Boston, Massachusetts, United States.
Fotini
F
Nicolaou
Retina Service, Angiogenesis Laboratory, Massachusetts Eye and Ear Infirmary, Department of Ophthalmology, Harvard Medical School, Boston, Massachusetts, United States.
Katarzyna
K
Brodowska
Retina Service, Angiogenesis Laboratory, Massachusetts Eye and Ear Infirmary, Department of Ophthalmology, Harvard Medical School, Boston, Massachusetts, United States.
Thanos D
TD
Papakostas
Retina Service, Angiogenesis Laboratory, Massachusetts Eye and Ear Infirmary, Department of Ophthalmology, Harvard Medical School, Boston, Massachusetts, United States.
Anna
A
Marmalidou
The Schepens Eye Research Institute, Massachusetts Eye and Ear, Department of Ophthalmology, Harvard Medical School, Boston, Massachusetts, United States.
Bruce R
BR
Ksander
Retina Service, Angiogenesis Laboratory, Massachusetts Eye and Ear Infirmary, Department of Ophthalmology, Harvard Medical School, Boston, Massachusetts, United States.
Joan W
JW
Miller
Retina Service, Angiogenesis Laboratory, Massachusetts Eye and Ear Infirmary, Department of Ophthalmology, Harvard Medical School, Boston, Massachusetts, United States.
Evangelos
E
Gragoudas
Retina Service, Angiogenesis Laboratory, Massachusetts Eye and Ear Infirmary, Department of Ophthalmology, Harvard Medical School, Boston, Massachusetts, United States.
Demetrios G
DG
Vavvas
D000620:Aminoimidazole Carboxamide / Q000031:analogs & derivatives* / Q000494:pharmacology; D015153:Blotting, Western; D002453:Cell Cycle / Q000187:drug effects; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D004789:Enzyme Activation / Q000187:drug effects*; D005434:Flow Cytometry; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D008545:Melanoma / Q000188:drug therapy / Q000201:enzymology / Q000235:genetics*; D017856:Phosphotransferases (Phosphate Group Acceptor) / Q000187:drug effects / Q000378:metabolism*; D012334:RNA, Neoplasm / Q000235:genetics*; D060888:Real-Time Polymerase Chain Reaction; D012265:Ribonucleotides / Q000494:pharmacology*; D058570:TOR Serine-Threonine Kinases / Q000235:genetics* / Q000378:metabolism; D014604:Uveal Neoplasms / Q000188:drug therapy / Q000201:enzymology / Q000235:genetics*; D000098943:Uveal Melanoma
D012334:RNA, Neoplasm; D012265:Ribonucleotides; D000620:Aminoimidazole Carboxamide; C546842:MTOR protein, human; D058570:TOR Serine-Threonine Kinases; C118888:AMP-dependent kinase (ATP-forming); D017856:Phosphotransferases (Phosphate Group Acceptor); C031143:AICA ribonucleotide
AICAR; AMPK; mTOR; melanoma