Engineered targeting tLyp-1 exosomes as gene therapy vectors for efficient delivery of siRNA into lung cancer cells

Abstract

Natural exosomes can express specific proteins and carbohydrate molecules on the surface and hence have demonstrated the great potentials for gene therapy of cancer. However, the use of natural exosomes is restricted by their low transfection efficiency. Here, we report a novel targeting tLyp-1 exosome by gene recombinant engineering for delivery of siRNA to cancer and cancer stem cells. To reach such a purpose, the engineered tLyp-1-lamp2b plasmids were constructed and amplified in Escherichia coli. The tLyp-1-lamp2b plasmids were further used to transfect HEK293T tool cells and the targeting tLyp-1 exosomes were isolated from secretion of the transfected HEK293T cells. Afterwards, the artificially synthesized siRNA was encapsulated into targeting tLyp-1 exosomes by electroporation technology. Finally, the targeting siRNA tLyp-1 exosomes were used to transfect cancer or cancer stem cells. Results showed that the engineered targeting tLyp-1 exosomes had a nanosized structure (approximately 100 nm) and high transfection efficiency into lung cancer and cancer stem cells. The function verifications demonstrated that the targeting siRNA tLyp-1 exosomes were able to knock-down the target gene of cancer cells and to reduce the stemness of cancer stem cells. In conclusion, the targeting tLyp-1 exosomes are successfully engineered, and can be used for gene therapy with a high transfection efficiency. Therefore, the engineered targeting tLyp-1 exosomes offer a promising gene delivery platform for future cancer therapy.

Engineering exosomes for targeted drug delivery

“…tLyP-1 peptide (CGNKRTR), a non-small cell lung cancer (NSCLC)-homing peptide, selectively targets neuropilin-1 (NRP1) and neuropilin-2 (NRP2) receptors in the cell membrane. tLyp-1 exosomes selectively delivered siRNA to human NSCLC cells 51 . In a recent report, we engineered chondrocyte affinity peptide (CAP, DWRVIIPPRPSA) exosomes that specifically delivered miRNA-140(miR-140)to chondrocytes in the joints and attenuated the progression of osteoarthritis (OA) in a rat model ( Figure 5 ) 52 .…”

Biodistribution of Exosomes and Engineering Strategies for Targeted Delivery of Therapeutic Exosomes

“…Click chemistry utilizes covalent interactions between an alkyne and azide residue to form a stable triazole linkage, which can be applied to attach targeting moieties on the surface of exosomes in a variety of aqueous buffers including water, alcohols, and dimethyl sulfoxide (DMSO) [84][85][86][87][88]. PEGylation, which is a modification of exosome's surface with branched polyethylene glycol (PEG), is one of a v integrin-targeting iRGD peptide Breast cancer cell line [110] NSCLC-homing peptide Tlyp-1 Lung cancer cell line [111] RVG peptide Brain/BBB [35] HER2 targeting DARPins HER2-expressing breast cancer [112] Conjugation with CD63…”

A smart viral vector for targeted delivery of hydrophobic drugs

“…tLyP-1, chosen as a targeting peptide, is an active CendR peptide that binds to neuropilin receptors (NRP1 and NRP2) that are typically overexpressed in the angiogenic vessels of most malignant tumor cells and in the majority of carcinomas 36 . So far, tLyP-1 has been conjugated to a variety of nano-delivery vehicles based on metal 37 , 38 , mesoporous silica 39 , PEG-PLA 40 , non-ionic surfactants 41 , liposomes 42 , 43 and exosomes 44 . tLyP-1 has also been conjugated with molecular trackers 36 , 45 , with other peptides 46 , and co-administered with drug-polymer conjugates 47 – 49 .…”