Plasma membrane nucleolin is a receptor for the anticancer aptamer AS1411 in MV4-11 leukemia cells.

Abstract

AS1411 is a DNA aptamer that is in phase II clinical trials for relapsed or refractory acute myeloid leukemia and for renal cell carcinoma. AS1411 binds to nucleolin, a protein that is overexpressed in the cytoplasm and on the plasma membrane of some tumor cells compared with normal cells. Studies were performed to determine whether cell surface nucleolin is a receptor for AS1411 in the acute myeloid leukemia cell line MV4-11. Biotinylation of MV4-11 cell surface proteins followed by immunoblotting of the biotinylated proteins showed that full-length (106 kDa) and truncated forms of nucleolin were present on the cell surface. In contrast, K-562 cells, which are 4-fold less sensitive than MV4-11 cells to AS1411, showed no full-length nucleolin and lesser amounts of the truncated forms of nucleolin on the cell surface. Incubation of MV4-11 cells with [(32)P]AS1411 and immunoprecipitation of the plasma membrane fraction with anti-nucleolin antibody demonstrated the presence of [(32)P]AS1411-nucleolin complexes. Anti-nucleolin antibody inhibited binding of fluorescein isothiocyanate (FITC)-AS1411 to plasma membrane nucleolin 56 +/- 10% SE (P < 0.01) compared with cells incubated with FITC-AS1411 only. Cellular uptake of [(32)P]AS1411 into MV4-11 cells was blocked by a 20-fold excess of unlabeled AS1411 but not by a 20-fold excess of the biologically inactive oligonucleotide CRO-26. Uptake was approximately 3-fold faster into MV4-11 cells than into K-562 cells. Partial knockdown of plasma membrane and cytosolic nucleolin in MCF-7 cells resulted in a 3-fold decrease in AS1411 uptake. These results provide evidence that plasma membrane nucleolin is a functional receptor for AS1411 in MV4-11 cells.

• Authors: Sridharan Soundararajan, Li Wang, Vijayalakshmi Sridharan, Weiwei Chen, Nigel Courtenay-Luck, David Jones, Eleanor K Spicer, Daniel J Fernandes
• Journal: Molecular pharmacology (Mol Pharmacol) Vol. 76 Issue 5 Pg. 984-91 (Nov 2009) ISSN: 1521-0111 [Electronic] United States
• PMID: 19657047 (Publication Type: Comparative Study, Journal Article, Research Support, N.I.H., Extramural, Research Support, Non-U.S. Gov't)

Chemical References

• AGRO 100
• Antineoplastic Agents
• Aptamers, Nucleotide
• Membrane Proteins
• Oligodeoxyribonucleotides
• Phosphoproteins
• RNA-Binding Proteins
• Receptors, Cell Surface
• nucleolin

Topics

• Animals
• Antineoplastic Agents (metabolism, pharmacology, therapeutic use)
• Aptamers, Nucleotide (metabolism, pharmacology, therapeutic use)
• Cell Line, Tumor
• Humans
• K562 Cells
• Leukemia, Myeloid, Acute (drug therapy, metabolism)
• Membrane Proteins (metabolism)
• Mice
• Oligodeoxyribonucleotides (metabolism, pharmacology, therapeutic use)
• Phosphoproteins (metabolism)
• Protein Binding (drug effects, physiology)
• RNA-Binding Proteins (metabolism)
• Receptors, Cell Surface (metabolism)