SiRNF8 Delivered by DNA Framework Nucleic Acid Effectively Sensitizes Chemotherapy in Colon Cancer

Zhao Guo,1,* Haoyun Song,1,* Yingxia Tian,2,* Jie Xu,3 Guokun Zhang,1 Yanan Guo,1 Rong Shen,1 Degui Wang1 1Department of Anatomy and Histology, Lanzhou University School of Basic Medical Sciences, Lanzhou, 730000, People’s Republic of China; 2Department of Internal Medicine, Gansu Provincial Academic Institute for Medical Research, Lanzhou, 730050, People’s Republic of China; 3Cuiying Biomedical Research Center, Lanzhou University Second Hospital, Lanzhou, Gansu, 730030, People’s Republic of China*These authors contributed equally to this workCorrespondence: Degui Wang; Rong Shen, Department of Anatomy and Histology, Lanzhou University School of Basic Medical Sciences, Lanzhou, 730000, People’s Republic of China, Tel +86 13893609120 ; +86 13909483373, Fax +86 931 8915023, Email wangdegui@lzu.edu.cn; shenr@lzu.edu.cnBackground: The evident side effects and decreased drug sensitivity significantly restrict the use of chemotherapy. However, nanoparticles based on biomaterials are anticipated to address this challenge.Methods: Through bioinformatics analysis and colon cancer samples, we initially investigated the expression level of RNF8 in colon cancer. Next, we constructed nanocarrier for delivering siRNF8 based on DNA tetrahedron (si-Tet), and Doxorubicin (DOX) was further intercalated into the DNA structure (si-DOX-Tet) for combination therapy. Further, the effects and mechanism of RNF8 inhibition on the sensitivity of colon cancer cells to DOX chemotherapy have also been studied.Results: RNF8 expression was increased in colon cancer. Agarose gel electrophoresis, transmission electron microscopy, and size distribution and potential analysis confirmed the successful preparation of the two nanoparticles, with particle sizes of 10.29 and 37.29 nm, respectively. Fluorescence imaging reveals that the carriers can be internalized into colon cancer cells and escape from lysosomes after 12 hours of treatment, effectively delivering siRNF8 and DOX. Importantly, Western blot analysis verified treatment with 50nM si-Tet silenced RNF8 expression by approximately 50% in colon cancer cells, and combined treatment significantly inhibited cell proliferation. Furthermore, the CCK-8 assay demonstrated that si-Tet treatment enhanced the sensitivity of colon cancer cells to the three chemotherapeutic drugs. Significant more DNA damage was detected after treatment with both si-Tet or si-DOX-Tet. Further flow cytometry analysis revealed that si-DOX-Tet treatment led to significantly more apoptosis, approximately 1.6-fold higher than treatment with DOX alone. Mechanistically, inhibiting RNF8 led to decreased ABCG2 expression and DOX efflux, but increased DNA damage, thereby enhancing the chemotherapeutic effect of DOX.Conclusion: We have successfully constructed si-DOX-Tet. By inhibiting the expression of RNF8, it enhances the chemotherapy sensitivity of DOX. Therefore, this tetrahedral FNA nanocarrier offers a new approach for the combined treatment of colon cancer. Keywords: DNA framework nucleic acid, RNF8, chemosensitivity, RNAi, colorectal cancer