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XIANG XU

China Pharmaceutical University
School of Pharmacy

Synchronous Conjugation of i-motif DNA and Therapeutic siRNA on the Vertexes of Tetrahedral DNA Nanocages for Efficient Gene Silence

Xiang Xu: TeamMember
Xiang Xu: Pro Gallery

Purpose(s)

1. Introducing exogenous nucleic acid to specifically repair abnormal gene expression at the molecular level.
2. Safe and effective delivery of small interfering RNA to inhibit the expression of epidermal growth factor receptor.
3. DNA tetrahedron nanocages transport siRNA into cancer cells efficiently via high transfection efficiency and escape lysosomal destruction.

Xiang Xu: Text

Method(s)

1. Linker strands were selected via structure prediction software and used to immobilize lysosomal-escaping overhang (X) strands.
2. PCR annealing technology was exploited to self-assemble the DNA strands into a regular tetrahedral nanocages in one step.
3. Confocal laser microscopy, Western Blot, and RT-qPCR respectively proved the lysosomal escape ability and the expression levels of EGFR protein and mRNA.
4. In vivo tumor inhibition experiment was performed on mice.

Xiang Xu: Text

Result(s)

In this study, we developed a series of DNA tetrahedron nanocages (Td), via synchronous conjugating different numbers of i-(X) and therapeutic siRNA on four vertexes of tetrahedral DNA nanocage (aX-Td@bsiRNA, a+b=4). This i-motif-conjugated Td exhibited good endosomal escape behaviors in A549 tumor cells, and the escape efficiency was affected by the number of i-motif. Furthermore, the downregulating mRNA and protein expression level of epidermal growth factor receptor (EGFR) caused by this siRNA embedded Td were verified in A549 cells. The tumor growth inhibition efficiency of the 2X-Td@2siRNA treated group in tumor-bearing mice was significantly higher than that of non-i-motif-conjugated Td@2siRNA (3.14-fold) and free siRNA (3.63-fold). These results demonstrate a general strategy for endowing DNA nanostructures with endosomal escape behaviours to achieve effective in vivo gene delivery and therapy.

Xiang Xu: Text

Conclusion(s)

In summary, a series of DNA nanostructures (aX-Td@bsiRNA) gifted with favorable endosomal escape behaviors were developed to achieve efficient siRNA in vivo delivery. This siRNA delivery system can supply the multifunctionality required to address challenges for siRNA in vivo delivery, including protecting siRNA from nuclease-mediated degradation, facilitating its cellular uptake, and promoting endosomal escape.
CLSM, Western Blot and in vivo anti-tumor results represent that DNA tetrahedrons modified with overhang as promising siRNAbased therapeutics carriers have achieved great progress in further in vivo delivery for gene therapy.

Xiang Xu: Text
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