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. 2009 Oct 23;38(1):e3. doi: 10.1093/nar/gkp841

Figure 1.

Figure 1.

Gymnotic delivery leads to long-term Bcl-2 silencing in 518 A2 melanoma cells. (A) Time course of Bcl-2 protein silencing in 518A2 melanoma cells after gymnotic delivery of SPC2996. Maximum protein silencing was observed after ∼6–7 days for SPC2996. C, untreated control. (B) RT–PCR evaluation of Bcl-2 mRNA levels in 518A2 melanoma cells demonstrating silencing after 6 days in the gymnotic delivery experiment in Figure 1A. C, untreated control; C*, single primer only. (C) Western blots in 518A2 melanoma cells demonstrating silencing of Bcl-2 protein by gymnotic delivery (6 days) of the sequence-specific anti-Bcl-2 phosphorothioate LNA gap-mer SPC2996. Three control oligomers with scrambled sequence, of which one (SPC3053) has identical chemistry to SPC2996, were inactive. (D) Iterative re-addition of SPC2996 (2.5 µM) produces long-term silencing. 518A2 melanoma cells were treated with SPC2996 at a plating density to ensure that the cells were confluent only by Day 10, and then re-plated at that density in the presence of 2.5 µM oligomer. This process was repeated after 10 additional days. Seventeen repetitions were performed. Bcl-2 protein silencing was continuously maintained after 180 consecutive days. However, if the oligomer was removed after 180 days, baseline levels of Bcl-2 protein were restored after 3 days.