Synergistic Treatment of Tumor by Targeted Biotherapy and Chemotherapy via Site-Specific Anchoring of Aptamers on DNA Nanotubes

Abstract

Run Chen,1,* Pengchao Sun,2,* Xiao Chu,1 Xiaorong Pu,1 Yang Yang,1 Nan Zhang,1,3,4 Yongxing Zhao1,3,4 1Department of Pharmaceutics, School of Pharmaceutical Sciences, Zhengzhou University, Zhengzhou, Henan 450001, People’s Republic of China; 2Institute for Biological Interfaces 1, Karlsruhe Institute of Technology, Karlsruhe 76344, Germany; 3HeNan Key Laboratory of Targeting Therapy and Diagnosis for Critical Diseases, Zhengzhou, Henan 450001, People’s Republic of China; 4Key Laboratory of Advanced Pharmaceutical Technology, Ministry of Education of China, Zhengzhou, Henan 450001, People’s Republic of China*These authors contributed equally to this workCorrespondence: Yongxing Zhao; Nan ZhangSchool of Pharmaceutical Sciences, Zhengzhou University, No. 100 KeXue Ave, Zhengzhou, HeNan 450001, People’s Republic of ChinaTel [phone]Fax [phone]Email [email protected]; [email protected]: Aptamers have been widely used as targeted therapeutic agents due to its relatively small physical size, flexible structure, high specificity, and selectivity. Aptamers functionalized nanomaterials, not only enhance the targeting of nanomaterials, but can also improve the stability of the aptamers. We developed aptamer C2NP (Apt) conjugated straight DNA nanotubes (S-DNT-Apt) and twisted DNA nanotubes (T-DNT-Apt) as nanocarriers for doxorubicin (DOX).Methods: The twisted DNA nanotubes (T-DNT) and straight DNA nanotubes (S-DNT) were assembled with a scaffold and hundreds of staples. Apt was site-specifically anchored on DNA nanotubes with either different spatial distribution (3 or 6 nm) or varied stoichiometry (15Apt or 30Apt). The developed nanocarriers were characterized with agarose gel electrophoresis and transmission electron microscopy. The drug loading and release in vitro were evaluated by measuring the fluorescence intensity of DOX using a microplate reader. The stability of DNT in cell culture medium plus 10% of FBS was evaluated by agarose gel electrophoresis. The cytotoxicity of DNA nanostructures against K299 cells was tested with a standard CCK8 method. Cellular uptake, cell apoptosis, cell cycle and reactive oxygen species level were investigated by flow cytometry. The expression of p53 was examined by Western Blot.Results: T-DNT-30Apt-6 exhibited the highest cytotoxicity when the concentration of Apt was 120 nM. After intercalation of DOX, the cytotoxicity of DOX@T-DNT-30Apt-6 was further enhanced due to the combination of chemotherapy of DOX and biotherapy of Apt. The enhanced cytotoxicity of DOX@T-DNT-30Apt-6 can be explained by the increase in the cellular uptake, cell apoptosis and intracellular ROS levels. Additionally, the interaction between Apt and its receptor CD30 could upregulate the expression of p53.Conclusion: These results demonstrate that both stoichiometry and spatial arrangement of Apt on T-DNT-Apt influence the anticancer activity. The developed twisted DNA nanotubes may be a solution for the synergistic treatment of cancer.Keywords: DNA nanotubes, aptamers, controlled spatial distribution, biotherapy and chemotherapy

Keywords

• dna nanotubes
• aptamers
• controlled spatial distribution
• biotherapy and chemotherapy