Calcium dependent cellular uptake of a c-myc antisense oligonucleotide.
Because a major limitation of ODN (oligodeoxynucleotide) use is inefficient cellular uptake, methods to improve ODN uptake could have important implications in the investigational and possibly therapeutic use of ODNs. In this study, antisense c-myc ODN cellular uptake in elevated extracellular calcium was increased up to 48-fold in the four cell lines examined. The role of calcium in ODN cellular uptake was examined using a 21-base ODN complementary to the c-myc proto-oncogene and the Rauscher cells. Cells were pretreated with uptake inhibitors in either 1.8 (physiologic) or 5.4 mM calcium prior to addition of (32P) labelled ODN. In physiologic calcium conditions, ODN cellular uptake was partially dependent on cellular energy and a trypsin-sensitive surface protein. In contrast, in the presence of elevated (5.4 mM) extracellular calcium, trypsinization and metabolic inhibition had a reduced and no effect, respectively, on uptake. Endocytosis and lysosomotropic inhibitors did not decrease uptake in either calcium concentrations. Therefore, the mechanism of ODN uptake may depend on the level of extracellular calcium. Furthermore, surface binding accounted for approximately 60% of total uptake in both physiologic and elevated calcium concentrations, suggesting that the increased uptake was not due exclusively to increased surface binding. Thus, the predominant mechanism of ODN uptake may depend on the extracellular calcium concentration.
Wu-Pong
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Department of Pharmaceutical Chemistry, University of California, San Francisco 94143.
Weiss
T L
TL
Hunt
C A
CA
D000818:Animals; D001152:Arsenicals / Q000494:pharmacology; D001483:Base Sequence; D001692:Biological Transport / Q000187:drug effects; D002118:Calcium / Q000494:pharmacology*; D002460:Cell Line; D002738:Chloroquine / Q000494:pharmacology; D003847:Deoxyglucose / Q000494:pharmacology; D016259:Genes, myc; D006801:Humans; D007700:Kinetics; D004915:Leukemia, Erythroblastic, Acute; D008264:Macrophages; D008969:Molecular Sequence Data; D008985:Monensin / Q000494:pharmacology; D016376:Oligonucleotides, Antisense / Q000378:metabolism*; D000090063:Proto-Oncogene Mas; D014357:Trypsin / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D001152:Arsenicals; C000715470:MAS1 protein, human; D016376:Oligonucleotides, Antisense; D000090063:Proto-Oncogene Mas; C029341:oxophenylarsine; D002738:Chloroquine; D008985:Monensin; D003847:Deoxyglucose; D014357:Trypsin; D002118:Calcium