Construction Of DNA Nanomaterials And Their Applications In Cell-related Research

The stability of genetic information and the diversity of living organisms were accurately maintained by DNA,the carrier of genetic information,via Watson-Crick base pairing.Meanwhile,various DNA nanomaterials can be constructed based on these precise base-pairing rules.DNA nanomaterials have attracted extensive attention in drug delivery,biosensing and cell surface engineering for they hold several advantages,such as ease of preparation and modification,high designability and high biocompatibility.While,there are still some challenges in applying DNA nanomaterials to cell-related research.First,due to the heterogeneity of cancer,different patients with the same type of cancer,or even the same patient at different disease stages,can result in totally different responses to cancer medications.Traditional targeted drug delivery systems are difficult to overcome cancer heterogeneity due to the inevitable off-target effects and the difficulty of achieving on-demand drug release in cells.The development of smart and customized targeted drug delivery systems that can achieve on-demand drug release in target cells is expected to overcome cancer heterogeneity and achieve personalized medicine,but still remains a challenge.Second,DNA nanomaterials has been modified onto cell membrane through several strategies and widely used in biosensing and biological regulation,while,there is still a need for methods that can graft DNA nanomaterials onto cell membrane with high speed,high efficiency,high stability,strong universality and good biocompatibility.Third,DNA has been used to regulate cell-cell interactions,while,the effects of interaction between DNAs and membrane-anchored ability of DNA on regulating cell-cell interactions remain to be studied.Meanwhile,cell-cell interactions are now regulated primarily by relatively static DNA hybridization or molecular recognition,while,regulating cell-cell interactions based on cell behavior will be more conducive to study cell biology.To address these issues,we first developed a smart drug delivery system to enable personalized medicine at the cellular level.Subsequently,we developed a new method for cell surface engineering.Finally,we applied DNA nanomaterials to regulate cell-cell interactions.The specific research contents of this thesis are as follows:(1)In Chapter 2,by using DNA self-assembly technology,we constructed a cell-customized drug delivery system at the cellular level for personalized medicine.The drug is a linear DNA duplex formed by tandem hybridization of two antisense oligonucleotide.And the two ends of the drug are blocked by aptamer AS1411 which can specifically bind to cancer cells and a molecular probe(ABP)that can sense and bind to a cancer marker,respectively.Our results indicated that under the guidance of AS1411,the DNA nanodrugs can be specifically delivered to cancer cells.Once c-raf-1 mRNA,a cancer marker,is detected,ABP can bind to it and thus activating the drug delivery systems.Then the activated drug delivery systems can induce cancer cell apoptosis by hybridizing with miRNA-21 and miRNA-150.Since the expression level of the cancer markers are related to the pathological state of the cells and the drug delivery system can be specifically activated by cancer markers,the drug delivery system can enable customized administration of individual cell and achieve personalized medicine at cellular level.(2)In Chapter 3,by using DNA self-assembly technology,we constructed amphiphilic DNA tetrahedral probes to develop a novel method for cell surface engineering.The amphiphilic DNA tetrahedral probes contain four vertices,one of which was modified by a functional nucleic acid and the other three vertices were modified by cholesterols.The amphiphilic DNA tetrahedral probes can be engineered to cell surface through the strong hydrophobic interaction between cholesterols and cell membrane.Our results indicated that compared with the traditional amphiphilic linear DNA probes,amphiphilic DNA tetrahedral probes can effectively improve the membrane-anchoring ability and biostability of functional nucleic acids.Combined the designability of DNA with the richness of chemical and biological activities of DNA,diverse functional DNA molecules can be engineered to cell surface through amphiphilic DNA tetrahedral probes,thus provide new methods for constructing sensing interfaces and engineering cell membrane with functional materials.(3)In chapter 4,we studied some basic properties of amphipathic DNA tetrahedral probes in regulating cell-cell interactions.Our results indicated that the rate,efficiency and extent of cell adhesion can be regulated by DNA.Meanwhile,the stability of cell-cell adhesion is related to the membrane-anchoring stability of DNA.Furthermore,manipulation of the physical contact between cells through DNA hybridization are beneficial for cell communications.(4)In Chapter 5,we constructed a dynamic DNA nanoprobe on cell surface for cell adaptive regulation of cell-cell interactions.An amphipathic DNA tetrahedral scaffold that contains an ATP-responsive probe were first constructed and engineered to cell membrane.Once exposed to external stimuli,cells can secrete ATP,which can activate the DNA probes on cell membrane.Then the activated DNA probes can initiate tandem hybridization of two DNA functional monomers,thereby introducing functional modules on cell membrane.Finally,cell-cell interactions can be regulated by the formed functional modules.