Figure 1.

Aptamer-Targeted Delivery of CD25 siRNAs to Activated CD8⁺ T Cells In Vitro

(A) CHO cells were co-transfected with 4-1BB-CD25 or 4-1BB-luciferase (4-1BB-luc) conjugates and with the ΨCHECK reporter plasmid containing short sequences corresponding to the murine CD25 siRNA target cloned into the 3′-untranslated region of Renilla luciferase. After 24 hr, the normalized Renilla luciferase activity was determined (Materials and Methods). (B) Purified CD8⁺ cells from C57BL/6 mice were polyclonally activated with a mixture of CD3 and CD28 antibodies and treated with conjugates three times at 500 nM concentration every 6 hr, starting 24 hr post activation. 24 hr after the last treatment, the cells were harvested and the levels of CD25 mRNA were assessed by qPCR. (C) 48 hr after the last treatment, the levels of CD25 protein on the cell surface were assessed by flow cytometry (n = 2). (D) CD25 expression on polyclonally activated CD8⁺ T cells incubated with 4-1BB or scrambled aptamer conjugated to CD25 siRNA (Scram-CD25).