Water-Soluble Cationic Polyphosphazenes Grafted with Cyclic Polyamine and Imidazole as an Effective Gene Delivery Vector.

Gene therapy holds immense potential as a future therapeutic strategy for the treatment of numerous genetic diseases which are incurable to date. Nevertheless, safe and efficient gene delivery remains the most challenging aspects of gene therapy. In this study, a series of polyphosphazenes (PPZ) bearing cyclic polyamine and imidazole groups were synthesized and investigated for gene delivery. Agarose gel electrophoresis assays showed that poly(imidazole/1,4,7,10-tetraazyclodocane)phosphazene (Im-PPZ-cyclen) had good binding ability with plasmid DNA (pDNA), yielding positively charged particles with a size around 120-140 nm from a ratio of 10:1 to 5:1 (Im-PPZ-cyclen/pDNA, w/w). The cytotoxicity of Im-PPZ-cyclen assayed by MTT was lower than that of PEI 25 kDa, and was similar to that reported for poly(di-2-dimethylaminoethylamine)phosphazene (poly(di-DMAEA)phosphazene) to some degree. The maximum transfection efficiency of Im-PPZ-cyclen/pDNA complexes against 293 T cells at the ratio of 5:1 (Im-PPZ-cyclen/pDNA, w/w) is close to that of Lipofectamine 2000. The present work may provide a strategy for the design of new cationic polymers with reduced cytotoxicity and be applied to gene delivery as an efficient nonviral vector.

Cyclen Grafted with poly[(Aspartic acid)-co-Lysine]: Preparation, Assembly with Plasmid DNA, and in Vitro Transfection Studies.

Development of safe and effective gene carriers is the key to the success of gene therapy. Nowadays, it is still required to develop new methods to improve nonviral gene delivery efficiency. Herein, copolymers of poly[(aspartic acid)-co-lysine] grafted with cyclen (cyclen-pAL) were designed and evaluated for efficient gene delivery. Two copolymers with different Asp/Lys block ratios were prepared and characterized by NMR and gel permeation chromatography analysis. Agarose gel retardation, circular dichroism, and fluorescent quenching assays showed the strong DNA-binding and protection ability for the title compounds. Atomic force microscopy studies clearly delineated uniform DNA globules with a diameter around 100 nm, induced by cyclen-pAL. By grafting cyclen on Asp, relatively high gene delivery efficiency and low cytotoxicity of the modified copolymers were achieved compared with their parent compounds. The present work might help to develop strategies for design and modification of polypeptide copolymers, which may also be applied to favorable gene expression and delivery.

Circular RNA AGAP1 Stimulates Immune Escape and Distant Metastasis in Renal Cell Carcinoma.

Clear cell renal cell carcinoma (ccRCC) is the most representative subtype of renal cancer, with a highly aggressive phenotype and extremely poor prognosis. Immune escape is one of the main reasons for ccRCC growth and metastasis, in which circular RNAs (circRNAs) play critical roles. Therefore, this research studied circAGAP1-associated mechanisms in immune escape and distant metastasis in ccRCC. circAGAP1/miR-216a-3p/MKNK2 was overexpressed or down-regulated by cell transfection. EdU assay, colony formation assay, scratch assay, Transwell assay, immunoblotting, and flow cytometry were used to evaluate cell proliferation, migration, invasion, EMT, and immune escape, respectively. Dual-luciferase reporting assay and RIP assay were used to evaluate the targeting relationship between circAGAP1/miR-216a-3p/MKNK2. Xenotransplantation in nude mice was used to evaluate the growth of ccRCC tumors in vivo. Here, circAGAP1 high expression was positively correlated with higher histological grade and distant metastasis and was a prognostic indicator for ccRCC. Depleting circAGAP1 effectively hampered the proliferative, invasive, and migratory capacities, EMT, and immune escape of ccRCC cells. Correspondingly, silencing circAGAP1 delayed tumor growth, distant metastasis, and immune escape in vivo. Mechanistically, circAGAP1 sponged the tumor suppressor miR-216a-3p, thereby preventing miR-216a-3p from inhibiting MAPK2. Collectively, our findings demonstrate that circAGAP1 exerts a tumor suppressor function through miR-216a-3p/MKNK2 during the immune escape and distant metastasis in ccRCC, and suggest that circAGAP1 may be a novel prognostic marker and therapeutic target for ccRCC.