Interleukin-6 receptor specific RNA aptamers for cargo delivery into target cells

Abstract

Aptamers represent an emerging strategy to deliver cargo molecules, including dyes, drugs, proteins or even genes, into specific target cells. Upon binding to specific cell surface receptors aptamers can be internalized, for example by macropinocytosis or receptor mediated endocytosis. Here we report the in vitro selection and characterization of RNA aptamers with high affinity (Kd = 20 nM) and specificity for the human IL-6 receptor (IL-6R). Importantly, these aptamers trigger uptake without compromising the interaction of IL-6R with its natural ligands the cytokine IL-6 and glycoprotein 130 (gp130). We further optimized the aptamers to obtain a shortened, only 19-nt RNA oligonucleotide retaining all necessary characteristics for high affinity and selective recognition of IL-6R on cell surfaces. Upon incubation with IL-6R presenting cells this aptamer was rapidly internalized. Importantly, we could use our aptamer, to deliver bulky cargos, exemplified by fluorescently labeled streptavidin, into IL-6R presenting cells, thereby setting the stage for an aptamer-mediated escort of drug molecules to diseased cell populations or tissues.

RAID3 - An interleukin-6 receptor-binding aptamer with post-selective modification-resistant affinity

“…These results indicate that areas favorable for RNA interaction on the surface of the IL-6R are not in close proximity to the interaction sites of the receptor responsible for IL-6 and gp130 binding. Combined with previous results obtained with aptamers like AIR-3 26,29 putatively targeting domain 1 of the IL-6R, our findings suggest that the structure of IL-6R does not favor the selection of aptamers directly interfering with signaling complex formation.…”

“…1B, column 2) may result from increased binding of the protein complex to the nitrocellulose membrane. A saturation of sIL-6R with the previously published aptamer AIR-3A 29 had no influence on F4 binding (Fig. 1B, column 6), confirming that F4 and AIR-3A bind to different sites on the sIL-6R.…”

“…26,29 Preliminary results indicate domain 1 of the IL-6R to be the dominant target for nucleic acid ligands in the selection process. This finding is in general agreement with experiences from other SELEX experiments and shows a bias for one target site in the selection of aptamers.…”

Aptamers as smart ligands for nano-carriers targeting

“…In comparison with antibodies, 98 aptamers exhibit enhanced potentials for application in diagnosis and treatment [27,28]. In 99 general, aptamers exhibit many advantages over antibodies and nanobodies which include simple 100 labeling procedure with various reporters for molecular recognition, ability of directional 101 amplification by PCR and low cost of production. Concurrently, aptamers exhibit the high 102 affinity and high degree of specificity for their targets comparable to monoclonal antibodies.…”

Structure and target interaction of a G-quadruplex RNA-aptamer

“…This 106 nt RNA aptamer could be truncated to its GQ forming binding motif AIR-3A (19 nt) and has recently been shown effective in targeted drug delivery. 28,29 While characterization of AIR-3(A) binding, cell internalization and therapeutic potential was successfully conducted, the knowledge of its structure is limited to the fact that it forms a parallel GQ 30 and that almost all nucleotides in the binding motif are essential to maintain affinity. 31 Here we report on the structural analysis performed on AIR-3 and AIR-3A.…”