Application Of Nucleic Acid Aptamer-mediated Docetaxel-carrying Albumin Nanoparticles In Colon Cancer Targeted Therapy

Objective:Colon cancer is a common malignant tumor that poses a great threat to human health.The incidence and mortality of colorectal cancer rank third among all malignant tumors.Because the symptoms of colon cancer are relatively insidious,many patients are already in the middle and advanced stages when they are diagnosed.Chemotherapy is the main treatment for patients with advanced colon cancer.However,traditional chemotherapy regimens are often accompanied by obvious side effects,which seriously reduce the quality of life of patients.Targeted therapy can selectively deliver cytotoxic drugs to tumor cells while reducing damage to normal tissue.Therefore,it has important application prospects in improving the efficacy of chemotherapy and reducing serious adverse effect caused by the drugs.Targeted therapy usually requires a targeted drug delivery system(TDDS),which often includes three components:tumor-targeting ligands,nanocarriers,and cytotoxic drugs.Nucleolin is usually found in the nucleus,but is also highly expressed on the cell surface of many tumors,including colon cancer.Therefore,nucleolin is a potentially important therapeutic target for targeted tumor therapy.Aptamer is a type of ligands made of single-stranded oligonucleotides.By forming complicated 3D structure,aptamer can bind to molecular targets with high specificity and affinity.Aptamer also has low immunogenicity,low production cost,and small molecular weight,with the advantages of good tumor tissue penetration.As a result,aptamer is regarded as a tumor-targeting ligand with great application potential.Albumin is an endogenous protein with low immunogenicity,and an excellent component for making biocompatible nanoparticles.Docetaxel(DTX)is a broad-spectrum anticancer drug,which is clinically used for treatment of a wide variety of tumors,such as cervical cancer,non-small cell lung cancer,and metastatic breast cancer.So far,however,aptamer-guided and DTX-loaded albumin nanoparticle has not been evaluated for colon cancer treatment.AS 1411 is a nucleolin-binding aptamer that has been approved for clinical trials.In this study,we plan to build a new TDDS with DTX-loaded albumin NP that is AS1411-modified(Apt-NPs-DTX).We plan to study Apt-NPs-DTX’s therapeutic effect on colon cancer with in vitro and in vivo experiments.Of note,AS 1411,albumin,and other components of Apt-NPs-DTX have been approved by US FDA for human applications.Methods:The 5’ end of the AS 1411 aptamer was modified with a sulfhydryl group,and was connected to the amino group on the albumin through the coupling agent SMCC.Through the self-assembly method,docetaxel was loaded into the albumin functionalized with AS1411 to form TDDS(Apt-NPs-DTX).The morphology of nanoparticles(NPs)was studied by transmission electron microscope.The particle size,polydispersity coefficient(PDI),and zeta potential of nanoparticles were evaluated by dynamic light scattering method.The encapsulation rate,drug loading rate,and drug release curve were studied by high performance liquid chromatography.Agarose gel electrophoresis was used to evaluate whether the aptamer was connected to the albumin NPs.The albumin NPs was loaded with fluorescent dyes,and flow cytometry and laser confocal microscopy were used to evaluate the targeted delivery of the load to CT26 colon cancer cells.The cellular anticancer efficacy of Apt-NPs-DTX was assayed by in vitro cytotoxicity experiments.In vivo animal experiments were conducted to evaluate the antitumor efficacy of Apt-NPs-DTX,its impact on survival,and side effects.Results:Apt-NPs-DTX has an average diameter of 62 nm and a zeta potential of-31.2 mV.DTX is released from albumin nanoparticles with a typical sustained-release profile.Compared with control cells,nucleolin-positive CT26 colon cancer cells preferentially ingest aptamer-mediated NPs.In vitro cytotoxicity studies shows that Apt-NPs-DTX significantly enhances the cytotoxicity against CT26 colon cancer cells.Animal experiments shows that,compared with non-targeted nanoparticles,Apt-NPs-DTX significantly improves the antitumor efficacy and prolonged the survival of tumor-bearing mice,without generating extra adverse effect.Conclusion:In this study,AS 1411-modified and docetaxel-loaded NPs are constructed for targeted treatment of colon cancer.The results show that Apt-NPs-DTX can selectively deliver docetaxel to colon cancer cells in vitro,and significantly enhance the antitumor efficacy in vivo,indicating that the TDDS has application potential in targeted therapy against colon cancer.