Design and synthesize of Folic Acid conjugated MOF-Chitosan nanoparticles containing LNA miRNA-224 antisense for targeted gene therapy of Colon Cancer

Design and synthesize of Folic Acid conjugated MOF-Chitosan nanoparticles containing LNA miRNA-224 antisense for targeted gene therapy of Colon Cancer
Negin Mokri,¹ Sepideh Khaleghi,^2,* Negar Motakef Kazemi,³ Mehrdad Hashemi,⁴
1. Department of Medical Biotechnology, Faculty of Advanced Sciences and Technology, Tehran Medical Sciences, Islamic Azad University, Tehran, Iran.
2. Department of Medical Biotechnology, Faculty of Advanced Sciences and Technology, Tehran Medical Sciences, Islamic Azad University, Tehran, Iran
3. Department of Medical Nanotechnology, Faculty of Advanced Sciences and Technology, Tehran Medical Sciences, Islamic Azad University, Tehran, Iran.
4. Department of Genetics, Faculty of Advanced Science and Technology, Tehran Medical Science, Islamic Azad University, Tehran, Iran.
Introduction: With the ability to control the expression of genes, miRNAs can be considered as one of the regulators of biological processes. Nanoparticles also have the ability to cause many factors at their level, including drugs, fluorescent dyes, and various ligands. Conjugate that all of these factors are effective in prolonging their survival in the blood and increasing the correct targeting of tumors . in this study, targeted MOF-Chitosan nanoparticles were synthesized with folic acid-containing miRNA-224 antibody sequence and were used to inhibit folic acid-receiving clone cancer cells. Thus, processes related to inhibiting cancer processes such as growth inhibition, cell death, apoptosis, and autophagy were examined in cellular and molecular forms
Methods: In this project, MOF nanoparticles of zinc acetate nanoparticles were first synthesized in dimensions of less than 200 nm and the quality of synthesis was evaluated by DLS, FTIR, and TEM methods. In addition, cell toxicity, apoptosis, and cell cycle inhibition were assessed by cellular and molecular methods such as MTT, Real-time PCR, Cell cycle arrest Flowcytometry, Flow cytometry annexin V. In this project, MOF nanoparticles were synthesized with chitosan coating and then conjugated with folic acid.
Results: TEM results show spherical morphology with the porous center and jelly margin of chitosan-folic acid, FTIR results prove the connection of covalent folic acid and chitosan and the creation of MOF networks. Cellular analyzes of these nanoparticles show the targeted toxicity of MCFM224 nanoparticles to HCT116 cancer cells. The cell death type was confirmed by apoptosis and cell cycle control tests. The molecular analyzes of the treated cells show high expression of BAX and Caspase 9 apoptosis genes and Beclin1 and mTORC1 autophagy genes, which indicate the induction of delayed and autophagic apoptosis in HCT116 cells. The cell death type was confirmed by apoptosis and cell cycle control tests. The molecular analyzes of the treated cells show high expression of BAX and Caspase 9 apoptosis genes and Beclin1 and mTORC1 autophagy genes, which indicate the induction of delayed autophagic and apoptosis in HCT116 cells. The toxicity of nanoparticles and the viability of cells were assessed by the MTT test, which indicated high mortality of MCFM224 nanoparticles in the HCT116 cell lines. Since HCT116 cells represent folic acid receptors, they showed significant mortality compared to normal CRL1831 intestinal cells, with molecular and cellular results confirming this. The results of flow cytometry of apoptosis performed using annexin v and pi indicate a high percentage of apoptosis in HCT116 cells treated with MCFM224 nanoparticles. In addition, the inhibition of the cell cycle confirms the cellulosic apoptosis of cells that were treated at the same time and at the same concentration and under the same conditions. An increase in the percentage of cells in the Sub-G1 phase indicates the induction of apoptosis and the planned death of treated cells.
Conclusion: According to the results of various studies in recent years, as well as the research conducted in this study, we can hope to treat cancer with the help of MOF-Chitosan nanoparticles and also combine it with microRNA antisense strand, which by conducting research and Further studies can achieve better results in the treatment of colon cancer.
Keywords: colorectal cancer, MiRNA224, LNA, gene therapy, nanoparticles