An Aptamer-Functionalized DNA Circuit to Establish an Artificial Interaction between T Cells and Cancer Cells.

Nongenetic strategies that enable control over the cell-cell interaction network would be highly desired, particularly in T cell-based cancer immunotherapy. In this work, we developed an aptamer-functionalized DNA circuit to modulate the interaction between T cells and cancer cells. This DNA circuit was composed of recognition-then-triggering and aggregation-then-activation modules. Upon recognizing target cancer cells, the triggering strand was released to induce aggregation of immune receptors on the T cell surface, leading to an enhancement of T cell activity for effective cancer eradication. Our results demonstrated the feasibility of this DNA circuit for promoting target cancer cell-directed stimulation of T cells, which, consequently, enhanced their killing effect on cancer cells. This DNA circuit, as a modular strategy to modulate intercellular interactions, could lead to a new paradigm for the development of nongenetic T cell-based immunotherapy.

Ligand Dilution Analysis Facilitates Aptamer Binding Characterization at the Single-Molecule Level.

Cell-specific aptamers offer a powerful tool to study membrane receptors at the single-molecule level. Most target receptors of aptamers are highly expressed on the cell surface, but difficult to analyze in situ because of dense distribution and fast velocity. Therefore, we herein propose a random sampling-based analysis strategy termed ligand dilution analysis (LDA) for easily implemented aptamer-based receptor study. Receptor density on the cell surface can be calculated based on a regression model. By using a synergistic ligand dilution design, colocalization and differentiation of aptamer and monoclonal antibody (mAb) binding on a single receptor can be realized. Once this is accomplished, precise binding site and detailed aptamer-receptor binding mode can be further determined using molecular docking and molecular dynamics simulation. The ligand dilution strategy also sets the stage for an aptamer-based dynamics analysis of two- and three-dimensional motion and fluctuation of highly expressed receptors on the live cell membrane.

A photochemically covalent lock stabilizes aptamer conformation and strengthens its performance for biomedicine.

Aptamers' vast conformation ensemble consisting of interconverting substates severely impairs their performance and applications in biomedicine. Therefore, developing new chemistries stabilizing aptamer conformation and exploring the conformation-performance relationship are highly desired. Herein, we developed an 8-methoxypsoralen-based photochemically covalent lock to stabilize aptamer conformation via crosslinking the inter-stranded thymine nucleotides at TpA sites. Systematical studies and molecular dynamics simulations were performed to explore the conformation-performance relationship of aptamers, revealing that conformation-stabilized aptamers displayed better ability to bind targets, adapt to physiological environment, resist macrophage uptake, prolong circulation half-life, accumulate in and penetrate into tumor than their counterparts. As expected, conformation-stabilized aptamers efficiently improved the therapeutic efficacy of aptamer-drug conjugation on tumor-bearing mice. Collectively, our study has developed a general, simple and economic strategy to stabilize aptamer conformation and shed light on the conformation-performance relationship of aptamers, laying a basis for promoting their basic researches and applications in biomedicine.

Discovery of small molecules binding to the normal conformation of prion by combining virtual screening and multiple biological activity evaluation methods.

Conformational conversion of the normal cellular prion protein, PrPC, into the misfolded isoform, PrPSc, is considered to be a central event in the development of fatal neurodegenerative diseases. Stabilization of prion protein at the normal cellular form (PrPC) with small molecules is a rational and efficient strategy for treatment of prion related diseases. However, few compounds have been identified as potent prion inhibitors by binding to the normal conformation of prion. In this work, to rational screening of inhibitors capable of stabilizing cellular form of prion protein, multiple approaches combining docking-based virtual screening, steady-state fluorescence quenching, surface plasmon resonance and thioflavin T fluorescence assay were used to discover new compounds interrupting PrPC to PrPSc conversion. Compound 3253-0207 that can bind to PrPC with micromolar affinity and inhibit prion fibrillation was identified from small molecule databases. Molecular dynamics simulation indicated that compound 3253-0207 can bind to the hotspot residues in the binding pocket composed by ß1, ß2 and α2, which are significant structure moieties in conversion from PrPC to PrPSc.