Construction and Biological Evaluation of a Novel Integrin ανβ3-Specific Carrier for Targeted siRNA Delivery In Vitro

Abstract

(1) Background: The great potential of RNA interference (RNAi)-based gene therapy is premised on the effective delivery of small interfering RNAs (siRNAs) to target tissues and cells. Hence, we aimed at developing and examining a novel integrin αvβ3-specific delivery carrier for targeted transfection of siRNA to malignant tumor cells; (2) Methods: Arginine-glycine-aspartate motif (RGD) was adopted as a tissue target for specific recognition of integrin αvβ3. To enable siRNA binding, a chimeric peptide was synthesized by adding nonamer arginine residues (9R) at the carboxy terminus of cyclic-RGD dimer, designated as c(RGD)2-9R. The efficiency of 9R peptide transferring siRNA was biologically evaluated in vitro by flow cytometry, confocal microscopy, and Western blot; (3) Results: An optimal 10:1 molar ratio of c(RGD)2-9R to siRNA was confirmed by the electrophoresis on agarose gels. Both the flow cytometry and confocal microscopy results testified that transfection of c(RGD)2-9R as an siRNA delivery carrier was obviously higher than the naked-siRNA group. The results of Western blot demonstrated that these 9R peptides were able to transduce siRNA to HepG2 cells in vitro, resulting in efficient gene silencing; and (4) Conclusion: The chimeric peptide of c(RGD)2-9R can be developed as an effective siRNA delivery carrier and shows potential as a new strategy for RNAi-based gene therapy.

Noninvasive imaging of c(RGD)2‐9R as a potential delivery carrier for transfection of siRNA in malignant tumors

“…The positively charged peptide c(RGD) 2 ‐9R could complex nucleic acids through electrostatic interaction and specifically recognize certain tumor cells with overexpressed integrin α ν β 3 by RGD sequence. The in vitro ability and efficiency of c(RGD) 2 ‐9R transferring siRNA and realizing gene knockdown have been confirmed in our previous study …”

“…Radiochemical purity of 99m Tc‐c(RGD) 2 ‐9R/siRNA under different conditions was >94% over 6 hours (Figure C). Moreover, the results of our earlier study have illustrated the effectiveness of c(RGD) 2 ‐9R in transporting siRNA into target cells in vitro …”

“…Moreover, the results of our earlier study have illustrated the effectiveness of c(RGD) 2 -9R in transporting siRNA into target cells in vitro. 14…”

“…The in vitro ability and efficiency of c(RGD) 2 -9R transferring siRNA and realizing gene knockdown have been confirmed in our previous study. 14 In addition, successful gene delivery requires detailed and comprehensive understanding of in vivo trafficking of the carriers. 15 Radionuclide tracing technology is considered to be the only approach that could be successfully translated from the laboratory to the clinical setting.…”

Special Issue “Synthesis and Applications of Oligonucleotide Conjugates”

“…Accordingly, the conjugates show promise for development of therapeutic agents exhibiting combined antisense/anti-oxidant and BNCT activities. The third paper on therapeutic oligonucleotides describes an integrin specific transfection agent for delivery of siRNA to tumor cells [ 7 ]. The agent was a chimeric peptide containing two cyclic structures that incorporated an Arg-Gly-Asp motif for integrin recognition and a cell penetrating arginine nonamer for enhancement of cellular uptake.…”