Surface DNA-scaffolds equip multifunctional biomaterials for precise immune cell modulation

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Synthetic biodegradable materials can provide important advantages in controlling the localization, stability and bioactivity of presented biomolecules in vivo to improve the efficacy and safety of immune cell therapies. The multi-model display of biomolecules, especially proteins, on synthetic material surfaces relies on chemical conjugation methods that are often inefficient.  Dr. Huang’s previous work published in Small revealed that functional groups presented on nanoparticle surfaces through hybridized oligonucleotide backbones (siRNA) have significantly higher efficiency of protein attachment than single-stranded backbones. This suggests that steric inhibition is an important factor limiting the surface conjugation of biomolecules, in addition to the drop in conjugation efficiency that occurs with the decay of functional groups and multi-step conjugation. DNA, a natural polymer with programmable sequence and unique hybridization driven by hydrogen bond formation, may overcome the above obstacles and be advantageous when used as scaffolds for surface assembly. Thus, we designed a synthetic short DNA hybridization-mediated “one-step” biomolecule assembly strategy for surfaces, before which biomolecules are pre-conjugated with DNA strands complementary to the scaffold DNA as shown in our article, just published on Nature Nanotechnology.

We achieved a high density of DNA scaffolds on polymeric particles made of poly(lactic-co-glycolic acid) (PLGA) through using PLGA-DNA amphiphiles as surfactants during the emulsion fabrication protocol. We were excited to observe that this DNA-hybridization guided strategy yielded fast and near surface-saturated loading of biomolecules including antigens and antibodies. The improved loading density of surface biomolecules was found to be necessary for presenting antigens to robustly activate synthetic Notch receptors engineered on human primary T cells, and may also facilitate other immune cell activation where ligand clustering is required. Through varying the DNA scaffold sequences and controlling their ratios, we obtained precise ratiometric control of multiple protein attachments – something difficult to achieve by other chemistries. The ratiometric control of costimulatory ligands (anti-CD3 and anti-CD28) on the particle surfaces plays essential roles in natural T cell activation and expansion. We also show the versatility of this strategy by fabricating particles of different sizes and compositions. This method can be applied to other biomaterial architectures (e.g. scaffolds and nonspherical particles) where fragile biomolecules need to be presented on surfaces.

The densely-packed DNA scaffolds and tethered payload offered protection from enzymatic degradation, providing an in vivo half-life of 3-4 days for surface payloads. Micron-sized particles injected intratumorally showed excellent retention for localized activation of AND-gate chimeric antigen receptor (CAR)-T cells. We believe that the true promise of this platform will be realized by using particles to present multiple signals within the tumor microenvironment.  Functionalized particles could kick-start tumor killing by localized activation of CAR-T cells. T cell activity within the tumor could be further augmented by co-functionalization with additional cargos, for instance, a cytokine, a costimulatory ligand, and a checkpoint inhibitor tailored to circumvent immune evasion mechanisms of a particular tumor type, bringing new opportunities to improve immune cell modulation.

Xiao Huang

Postdoctoral researcher, University of California San Francisco