Cellular Uptake of Oligodeoxyribonucleoside Methylphosphonates

Abstract

The cellular uptake of oligodeoxyribonucleoside methylphosphonates has been evaluated using three radiolabeled oligomers. Oligomers I and II ([³H]-T₈ and [³H]-T₁₆, respectively) are nonionic methylphosphonate oligomers labeled with tritium on the phosphonate internucleotide linkage. EDA-III contains a single phosphodiester linkage, a [³²P]-label and an ethylenediamine conjugate at the [³²P]-5′-end. All three oligomers are stable in cells. At a 1 μM concentration, oligomer I is not taken up by human erythrocytes. The octanol/DPBS partition coefficients for oligomers I and II (1.5 x 10^-4 and 4.2 x 10^-4, respectively) further indicate that these molecules should not diffuse across cell membranes at appreciable rates. Oligomer I is taken up by HL-60 cells, although at a slower rate than the uptake of the fluid-phase marker sucrose. The cell-associated levels of oligomer II in K-562 cells following incubation of cells with the oligomer for 2 days is independent of concentration and nonsaturable, suggesting a mechanism of uptake independent of receptor. Finally, the initial uptake rate of EDA-III in mouse L cells is greater than the uptake of two oligodeoxyribonucleotides (T₈, T₁₆), reaching a plateau after 3 hours incubation with cells. These observations should aid in the elucidation of the mechanism by which this class of antisense agents enters the intracellular environment.