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Abstract
Background:
In vitro transcribed (IVT) mRNA has been applied as an alternative therapeutic molecule to plasmid DNA in the field of cancer therapy and biomedical research studies. mRNA-based therapy has demonstrated several advantages over its DNA counterparts. However, its further therapeutic application is largely restricted by delivery method.
Methods:
In this work, a liposome-protamine lipoplex (CLPP) was prepared to deliver IVT mRNA encoding survivin-T34A gene, forming a novel core-shell structured nanoparticle formulation (CLPP/mSur-T34A).
Results:
The prepared CLPP/mSur-T34A particle had an average size of 186.1±3.1 nm, displaying high mRNA transfecting and expression efficiency on C26 tumor cells through lipid rafts-mediated endocytosis. CLPP/mSur-T34A mRNA formulation demonstrated obvious therapeutic effects on various models of C26 colon cancer both in vitro and in vivo. Particularly, local and systemic administration of CLPP/mSur-T34A particle exhibited superior antitumor effect regarding its DNA plasmid counterpart with high safety.
Conclusion:
Our results indicated the high delivery capacity of liposome-protamine lipoplex and further suggested CLPP/mSur-T34A mRNA formulation to be a potential candidate for colon cancer therapy.
Acknowledgments
This work was supported by the National Major Scientific and Technological Special Project for “Significant New Drugs Development” (No. 2018ZX09733001), the National Key Specialty Construction Project of Clinical Pharmacy (No. 30305030698) and the Science Foundation of Chengdu (2016-HM01-00438-SF).
Abbreviations list
CLP, liposome; CLPP, liposome-protamine lipoplex; mSur-T34A, survivin-T34A mRNA; pSur-T34A, survivin-T34A plasmid DNA; mEGFP, EGFP mRNA; pEGFP, EGFP plasmid DNA; Lipo2K, LipofectamineTM2000; i.t., intratumoral injection; i.p., intraperitoneal injection; IVT, in vitro transcribed; IL-6, interleukin-6; TNF-α, tumor necrosis factor-α; IFN-γ, interferon-γ.
Disclosure
The authors report no conflicts of interest in this work.
Supplementary materials
Figure S1 Primer sequences.
Figure S2 Comparison of plasmid-based gene delivery by different vectors on C26 cells. (A) Images of transfected cells with different materials. (B) Transfection efficiencies calculated by flow cytometry.
Abbreviation: Fluor, fluorescence.
Figure S3 CLPP/mSur-T34A particles efficiently inhibited the growth of C26 cancer cells in vitro. (A) Evaluation of cell growth inhibition by CLPP/mSur-T34A particles after 48 hrs. (B) The change of cell morphology in each group. (C and D) 48 hrs post transfection, CLPP-delivered survivin-T34A mRNA efficiently induced apoptosis in C26 cells. (E and F) In abdominal cavity metastases, treatment with CLPP/mSur-T34A particles increased the mRNA level of caspase8 and caspase10 in tumor tissue. (G) In subcutaneous xenograft model, treatment with CLPP/mSur-T34A particles increased the mRNA level of caspase8 in tumor tissue. All data are reported as mean ± SEM and ***P<0.001, **P<0.01, *P<0.05 (two-tailed Student’s t-test).
Abbreviation: NS, normal saline.
Figure S4 HE analysis of main organs from each treatment group, no obvious pathological changes were observed (magnification =200×).
