Research And Biological Application Of Receptor Ligand Multivalent Interaction Based On Framework Nucleic Acids

Receptor-ligand interaction plays a key role in many physiological processes such as cell proliferation,signal transduction and immune activation.The ligands and receptors on the cell surface usually display the multivalent interactions.As a kind of DNA nanoscaffolds with precise conformation,the framework nucleic acids provide a reliable tool for the construction of multivalent ligands in vitro for revealing the mechanism of receptor-ligands interaction.By the nanoscale of framework nucleic acids,the number,spacing and spatial conformation of ligands can be precisely regulated.The binding characteristics and influencing factors of receptor ligand interaction on cell surface can be studied.Efficient molecular recognition and targeted therapy are realized with the optimized multivalent ligands.In this paper,we designed and constructed different multivalent ligands using framework nucleic acids.By regulating the valence,conformation,species and direction of the ligands,the affinity between ligands and receptor molecules were optimized,which can be applied to the study of cell pattern recognition imaging and targeted drug delivery.The specific research contents are as follows:(1)Directing multivalent aptamer-receptor binding on the cell surface with programmable atom-like nanoparticlesLigand direction is crucial for cell surface receptor ligand multivalent interactions.But the current multivalent ligand construction systems generally lack controllable programming for ligand direction.We designed programmable atom-like nanoparticles(apt PANs)to program the binding of multivalent aptamers to cell surface receptors.Tetrahedral DNA framework regulates the position of aptamers to realize the orthogonal atom-like valence and orientation,constructing multivalent aptamers with fixed valence but different direction.The apt PANs with certain flexibility and orientation showed adaptive binding to cell surface receptors.We demonstrate that apt PANs based on linear oligomer bind to tumor cells with high affinity(13 times stronger than monovalent aptamer)and inhibit cell growth by antagonizing receptor molecules on tumor cells(～50% inhibition ratio).(2)Precise conformation regulation of the antibody complementary determining region on nanoscale with framework nucleic acidsThe conformation of the complementary determination domain(CDR)in antibody determines the affinity and specificity of antigen-antibody interaction.However,the the conformation of CDR cannot be accurately controlled with current methods.So the effect of subtle conformational changes of CDR on the binding specificity and affinity of antibodies remained unclear.We used the addressability at atomic resolution of the framework nucleic acids to regulate the CDR conformation in the 10 nm range through chemical coupling of CDR peptides.Combined with molecular dynamics simulation,we reconstructed the active conformation of CDR and revealed the relationship between the CDR conformation and antibody affinity.(3)Cell imaging with multi-color DNA framework probesCell recognition based on cell surface biomarkers plays an important role in cancer diagnosis and targeted therapy.However,many biomarker expressions are not cellspecific,resulting in the lack of one-to-one specificity of recognition ligands,which limits the accuracy of cell recognition.We arranged different aptamers on fractal DNA frameworks to improve the dimension of aptamer-based cell recognition,and construct multi-color probes by encoding different fluorescence molecules to realize multiplex cell imaging and classification.(4)DNA origami-enabled engineering of ligand-drug conjugates for targeted drug deliveryTargeted drug delivery into tumor cells remains a challenge in the construction of drug delivery platforms.We designed a ligand-conjugated DNA origami nanostructure(DON)as a drug delivery platform for prostate cancer targeted therapy.Doxorubicin(Dox)is loaded into DON by intercalating into double-stranded DNA.This platform is capable of regulating the alignment of ligands with high efficiency of drug loading.Dox could be specifically delivered to PSMA positive cancer cell lines LNCa P and the therapeutic effect is related to the valence of ligands conjugated to DON.