Deacetylated-poly-N-acetylglucosamine-folic Acid as a Nanocarrier for Delivering miR-196a Inhibitor to Anticancer Activity

Background: MiR-196a is particularly noticeable in the development of liver cancer. However, the rapid degradation by ribonuclease (RNase) imposes a limit on the miRNA gene therapy applications. Aims: To design a novel gene-targeting nano system for liver cancer treatment. Study Design: Cell culture study and animal experimentation Methods: Deacetylated (DEAC)-poly-N-acetylglucosamine (PNAG)- folic acid (FA) was prepared via ethyl (dimethylaminopropyl) carbodiimide/N-hydroxysuccinimide reaction, and miR-196a inhibitor (miR-196a I)/DEAC-PNAG-FA was prepared through self-assembly. The characterization and nucleic acid protection of the self-assembly system were also determined. The biological function and related mechanism of the prepared system were studied at cellular and molecular levels. Mice were established as a xenotransplantation model to evaluate the anticancer capacity of miR-196a I/DEAC PNAG-FA in vivo. Results: The morphology of miR-196a I/DEAC-PNAG-FA was uniform, and its particle size was approximately 70–100 nm. A nanocarrier with an N/P ratio of 200:1 can maximize the nucleic acid carrying capacity of the self-assembly system. The nanosystem can protect miRNA from RNase degradation and could be internalized rapidly within 4 h. The self-assembly system significantly enhanced the apoptosis-inducing effect of miR-196a I on HepG2 cells (P = 0.003). Molecular biological analyses confirmed that the apoptosis inducing effect of the nanosystem was due to the inhibition of miR 196a gene expression in HepG2 cells, which upregulate the expression of pro-apoptotic proteins FOXO1 (P < 0.001), Bax (P < 0.001), Ki67 (P < 0.001), and proliferating cell nuclear antigen (P < 0.001), and inhibit the expression of apoptosis inhibitory protein Bcl-2 (P < 0.001). Moreover, compared with free miR-196a inhibitor or miR-196a I/ DEAC-PNAG, miR-196a I/DEAC-PNAG-FA can more effectively inhibit tumor growth in vivo (P = 0.026). Conclusion: The newly prepared self-assembly targeting system can effectively induce apoptosis and abrogate tumor growth, which may open a new approach for liver cancer treatment.

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1. Huang C, Zhang H, Bai R. Advances in ultrasound-targeted microbubble mediated gene therapy for liver fibrosis. Acta Pharm Sin B. 2017;7:447-452. [CrossRef]
2. Xiong Y, Fang JH, Yun JP, et al. Effects of microRNA-29 on apoptosis, tumorigenicity, and prognosis of hepatocellular carcinoma. Hepatology. 2010;51:836-845. [CrossRef]
3. Chen Y, Gao DY, Huang L. In vivo delivery of miRNAs for cancer therapy: challenges and strategies. Adv Drug Deliver Rev. 2015;81:128-141. [CrossRef]
4. Slezak-Prochazka I, Durmus S, Kroesen BJ, van den Berg A. MicroRNAs, macrocontrol: regulation of miRNA processing. RNA. 2010;6:1087-1095. [CrossRef]
5. Wan WJ, Qu CX, Zhou YJ, et al. Doxorubicin and siRNA-PD-L1 co-delivery with T7 modified ROS-sensitive nanoparticles for tumor chemoimmunotherapy. Int J Pharm. 2019;566:731-744. [CrossRef]
6. Kubota K, Onishi K, Sawaki K, et al. Effect of the nanoformulation of siRNA-lipid assemblies on their cellular uptake and immune stimulation. Int J Nanomedicine. 2017;12:5121-5133. [CrossRef]
7. Soliman C, Walduck AK, Yuriev E, et al. Structural basis for antibody targeting of the broadly expressed microbial polysaccharide poly-N-acetylglucosamine. J Biol Chem. 2018;293:5079-5089. [CrossRef]
8. Luo L J, Huang CC, Chen HC, Lai JY, Matsusaki M. Effect of deacetylation degree on controlled pilocarpine release from injectable chitosan-g-poly (N-isopropylacrylamide) carriers. Carbohyd Polym. 2018;197:375-384. [CrossRef]
9. Erdoğar N, Esendağlı G, Nielsen TT, et al. Therapeutic efficacy of folate receptor targeted amphiphilic cyclodextrin nanoparticles as a novel vehicle for paclitaxel delivery in breast cancer. J Drug Target. 2018;26:66-74. [CrossRef]
10. Liu MC, Liu L, Wang XR, et al. Folate receptor-targeted liposomes loaded with a diacid metabolite of norcantharidin enhance antitumor potency for H22 hepatocellular carcinoma both in vitro and in vivo. Int J Nanomedicine. 2016;11:1395-1412. [CrossRef]
11. Mirrahimi M, Hosseini V, Shakeri-Zadeh A, et al. Modulation of cancer cells’ radiation response in the presence of folate conjugated Au@ Fe2 O 3 nanocomplex as a targeted radiosensitizer. Clin Transl Oncol. 2019;21:479-488. [CrossRef]
12. Khademi S, Sarkar S, Shakeri-Zadeh A, et al. Corrigendum to “Targeted gold nanoparticles enable molecular CT imaging of head and neck cancer: An in vivo study” [Int. J. Biochem. Cell Biol. 114 (2019) 105554]. Int J Biochem Cell Biol. 2019;120:105695. [CrossRef]
13. Lee KY, Seow E, Zhang Y, Lim YC. Targeting CCL21-folic acid-upconversion nanoparticles conjugates to folate receptor-α expressing tumor cells in an endothelial tumor cell bilayer model. Biomaterials. 2013;34:4860-4871. [CrossRef]
14. Movahedi MM, Mehdizadeh A, Koosha F, et al. Investigating the photo-thermo radiosensitization effects of folate-conjugated gold nanorods on KB nasopharyngeal carcinoma cells. Photodiagn Photodyn. 2018;24:324-331. [CrossRef]
15. Lu CY, Chen SY, Peng HL, Kan PY, Chang WC, Yen C. Cell-free methylation markers with diagnostic and prognostic potential in hepatocellular carcinoma. Oncotarget. 2017;8:6406-6418. [CrossRef]
16. Hao YX, Wang JP, Zhao LF. Associations between three common MicroRNA polymorphisms and hepatocellular carcinoma risk in Chinese. Asian Pac J Cancer Prev. 2013;14:6601-6604. [CrossRef]
17. Darda L, Hakami F, Morgan R, Murdoch C, Lambert DW, Hunter KD. The role of HOXB9 and miR-196a in head and neck squamous cell carcinoma. PloS One. 2015;10:e0122285. [CrossRef]
18. Yang L, Peng F, Qin J, Zhou H, Wang B. Downregulation of microRNA-196a inhibits human liver cancer cell proliferation and invasion by targeting FOXO1. Oncol Rep. 2017;38:2148-2154. [CrossRef]
19. van der Ven CF, Wu PJ, Tibbitt MW, et al. In vitro 3D model and miRNA drug delivery to target calcific aortic valve disease. Clin Sci. 2017;131:181-195. [CrossRef]
20. Yu M, Xue Y, Ma PX, Mao C, Lei B. Intrinsic ultrahigh drug/miRNA loading capacity of biodegradable bioactive glass nanoparticles toward highly efficient pharmaceutical delivery. ACS Appl Mater Interfaces. 2017;9:8460-8470. [CrossRef]
21. Malik S, Lim J, Slack FJ, Braddock DT, Bahal R. Next generation miRNA inhibition using short anti-seed PNAs encapsulated in PLGA nanoparticles. J Control Release. 2020;327:406-419. [CrossRef]
22. Endo-Takahashi Y, Negishi Y, Nakamura A, et al. Systemic delivery of miR-126 by miRNA-loaded Bubble liposomes for the treatment of hindlimb ischemia. Sci Rep. 2014;4:3883. [CrossRef]
23. Robertson NM, Toscano AE, LaMantia VE, et al, et al. Unlocked Nucleic Acids for miRNA detection using two-dimensional nano-graphene oxide. Biosens Bioelectron. 2017;89:551-557. [CrossRef]
24. Fan Z, Zhu P, Zhu Y, Wu K, Li CY, Cheng H. Engineering long-circulating nanomaterial delivery systems. Curr Opin Biotech. 2020;66:131-139. [CrossRef]
25. El-Khatib AM, Badawi MS, Ghatass ZF, Mohamed MM, Elkhatib M. Synthesize of silver nanoparticles by arc discharge method using two different rotational electrode shapes. J Clust Sci. 2018;29:1169-1175. [CrossRef]
26. Vergote I, Leamon CP. Vintafolide: a novel targeted therapy for the treatment of folate receptor expressing tumors. Ther Adv Med Oncol. 2015;7:206-218. [CrossRef]
27. Koirala N, Das D, Fayazzadeh E, et al. Folic acid conjugated polymeric drug delivery vehicle for targeted cancer detection in hepatocellular carcinoma. J Biomed Mater Res A. 2019;107:2522-2535. [CrossRef]
28. Fernández M, Javaid F, Chudasama V. Advances in targeting the folate receptor in the treatment/imaging of cancers. Chem Sci. 2018;9:790-810. [CrossRef]
29. Jones Buie JN, Zhou Y, Goodwin AJ, et al. Application of Deacetylated Poly-N- Acetyl Glucosamine Nanoparticles for the Delivery of miR-126 for the Treatment of Cecal Ligation and Puncture-Induced Sepsis. Inflammation. 2019;42:170-184. [CrossRef]
30. Pu M, Chen J, Tao Z, et al. Regulatory network of miRNA on its target: coordination between transcriptional and post-transcriptional regulation of gene expression. Cell Mol Life Sci. 2019;76:441-451. [CrossRef]
31. Schanen BC, Li X. Transcriptional regulation of mammalian miRNA genes. Genomics. 2011;97:1-6. [CrossRef]
32. Lambert MP, Terrone S, Giraud G, et al. The RNA helicase DDX17 controls the transcriptional activity of REST and the expression of proneural microRNAs in neuronal differentiation. Nucleic Acids Res. 2018;46:7686-7700. [CrossRef]