Combined Self-Assembled Hendeca-Arginine Nanocarriers for Effective Targeted Gene Delivery to Bladder Cancer

Pu Zhang1 *, Haibao Zhang2 *, Bin Zheng1 *, Heng Wang,1 Xiaolong Qi,1 Shuai Wang,1 Zhenghong Liu,1 Li Sun,1 Yang Liu,3 Xiaowen Qin,1 Weijiao Fan,1 Minghai Ma,2 Wing-Fu Lai,1 Dahong Zhang1 1Urology & Nephrology Center, Department of Urology, Zhejiang Provincial People’s Hospital, Affiliated People’s Hospital, Hangzhou Medical College, Hangzhou, Zhejiang People’s Republic of China; 2Key Laboratory of Environment and Genes Related to Diseases, Xi’an Jiaotong University, Xi’an, People’s Republic of China; 3Cancer Center, Department of Ultrasound Medicine, Zhejiang Provincial People’s Hospital, Affiliated People’s Hospital, Hangzhou Medical College, Hangzhou, Zhejiang People’s Republic of China*These authors contributed equally to this workCorrespondence: Dahong Zhang; Wing-Fu Lai, No. 138 Shang Tang Road, Hangzhou, 310014, People’s Republic of China, Tel +86-13705711671, Email zppurology@163.com; rori0610@graduate.hku.hkIntroduction: Bladder cancer (BCa) is among the most prevalent cancers worldwide. However, the effectiveness of intravesical therapy for BCa is limited due to the short dwell time and the presence of the permeation barrier.Methods: Nanocomplexes were self-assembled between DNA and hendeca-arginine peptide (R11). Stepwise intravesical instillation of R11 and the generated nanocomplexes significantly enhanced the targeting capacity and penetration efficiency in BCa therapy. The involved mechanism of cellular uptake and penetration of the nanocomplexes was determined. The therapeutic effect of the nanocomplexes was verified preclinically in murine orthotopic BCa models.Results: Nanocomplexes exhibited the best BCa targeting efficiency at a nitrogen-to-phosphate (NP) ratio of 5 but showed a lack of stability during cellular uptake. The method of stepwise intravesical instillation not only increased the stability and target specificity of the DNA component but also caused the delivered DNA to more effectively penetrate into the glycosaminoglycan layer and plasma membrane. The method promotes the accumulation of the delivered DNA in the clathrin-independent endocytosis pathway, directs the intracellular trafficking of the delivered DNA to nonlysosome-localized regions, and enables the intercellular transport of the delivered DNA via a direct transfer mechanism. In preclinical trials, our stepwise method was shown to remarkably enhance the targeting and penetration efficiency of DNA in murine orthotopic BCa models.Conclusion: With this method, a stepwise intravesical instillation of self-assembled nanocomplexes, which are generated from hendeca-arginine peptides, was achieved; thus, this method offers an effective strategy to deliver DNA to target and penetrate BCa cells during gene therapy and warrants further development for future intravesical gene therapy in the clinical context.Graphical Abstract: Keywords: bladder cancer, intravesical therapy, DNA delivery, direct transfer