Periodic Reporting for period 3 - InActioN (Intracellular Action of DNA-based Nano-materials)
Periodo di rendicontazione: 2024-02-01 al 2025-07-31
Intracellular Action of DNA-based Nanomaterials
In the InActioN project, we aim to use DNA-based nanomaterials inside cells, in order to investigate molecular mechanisms within our immune system. Using DNA as a synthetic material offers the unique opportunity to precisely control the spacing of functional molecules like proteins or antibodies. One of the main advantages of using DNA as a material platform is that the components are encoded by natural DNA base pairing, with the result that all architectures are automatically perfect clones of one another. This uniformity is critical when studying interactions with biology, especially when we want to investigate how the number of interactions, or the geometric pattern how these interactions are made, play a role in the biological outcome. These questions can help us to gain a deeper understanding of how materials can interact with biology and how to better design therapeutics, diagnostics or even vaccines.
A critical requirement in order to interact with cellular mechanisms inside the cell, is a stable and controlled intracellular delivery of these nanomaterials. Naturally, DNA is found only in a cell nucleus, and cells have mechanisms in place to destroy what they see as foreign DNA when it is found outside the nucleus. Previous work has focused on the development of protective coatings, however, the impact of this coating on cell uptake had not been carefully analyzed. We are systematically increasing our fundamental understanding of DNA-cell interactions, and utilising this progress to engineer smart solutions to precise communication with natural processes inside the cell.
Our research encourages a shift in materials engineering, by showing that the geometry and flexibility of a material at the nanoscale can have a significant impact on the selectivity of its interactions. While we already demonstrated that uniform, precisely engineered materials can be used to provoke a specific immune response, this project holds a much wider relevance. Engineering a balance of geometry and flexibility for selective interactions is relevant for all molecular interfaces, from crystal self-assembly to biological function. As such, the fundamental guidelines of selective interactions at interfaces opens up a whole new platform of material design for nano-technology.
In the InActioN project, we aim to use DNA-based nanomaterials inside cells, in order to investigate molecular mechanisms within our immune system. Using DNA as a synthetic material offers the unique opportunity to precisely control the spacing of functional molecules like proteins or antibodies. One of the main advantages of using DNA as a material platform is that the components are encoded by natural DNA base pairing, with the result that all architectures are automatically perfect clones of one another. This uniformity is critical when studying interactions with biology, especially when we want to investigate how the number of interactions, or the geometric pattern how these interactions are made, play a role in the biological outcome. These questions can help us to gain a deeper understanding of how materials can interact with biology and how to better design therapeutics, diagnostics or even vaccines.
A critical requirement in order to interact with cellular mechanisms inside the cell, is a stable and controlled intracellular delivery of these nanomaterials. Naturally, DNA is found only in a cell nucleus, and cells have mechanisms in place to destroy what they see as foreign DNA when it is found outside the nucleus. Previous work has focused on the development of protective coatings, however, the impact of this coating on cell uptake had not been carefully analyzed. We are systematically increasing our fundamental understanding of DNA-cell interactions, and utilising this progress to engineer smart solutions to precise communication with natural processes inside the cell.
Our research encourages a shift in materials engineering, by showing that the geometry and flexibility of a material at the nanoscale can have a significant impact on the selectivity of its interactions. While we already demonstrated that uniform, precisely engineered materials can be used to provoke a specific immune response, this project holds a much wider relevance. Engineering a balance of geometry and flexibility for selective interactions is relevant for all molecular interfaces, from crystal self-assembly to biological function. As such, the fundamental guidelines of selective interactions at interfaces opens up a whole new platform of material design for nano-technology.
In the first phases of InActioN, we explored in detail the effect that different polymer coatings, as well as functionalization and labeling surface modifications, had on cell uptake efficiency. We identified coating-specific protein-coronea, and analyzed the downstream effect on uptake. Additionally, we have compiled a clear set of design parameters to guide the engineering or stable, yet potent, nanomaterials to enhance uptake.
Secondly, we have used these design guidelines to engineer materials to explore the importance of controlling the nano-spacing of vaccine adjuvants for selective activation of immune cells. Here, we coupled knowledge of pre-existing crystallography data of immune receptors to guide the design of spatially controlled ligand arrays on DNA-based nanomaterials. We created nanoparticles presenting CpG adjuvants to activate Toll-like Receptor 9 (TLR9) in a rationally designed way. Nanoscale variations in ligand spacing were shown to produce significantly different immunological responses: by matching CpG spacing to the distance of the TLR9 active dimer, cellular activation was significantly enhanced compared to random ligand presentation or free form CpG. For the first time, we show the tight connection between geometry and control of cellular signaling for CpG adjuvants. These findings are fundamental not only for a fine-tuned manipulation of the immune system, but also show a need to consider the importance of spatially controlled presentation and nanorigidity of therapeutics to increase efficacy and specificity of immune-modulating nanomaterials where multivalent binding interactions are involved.
We are currently exploring in more depth the potential avenues to use stabilizing coatings as transporter of our DNA nanomaterials into precise locations in the cytoplasm, as this is a rich pool of DNA-binding proteins. We aim to generate a toolbox that allows for an a-priori design of the best combination of coating / geometry / stability / uptake for a desired intracellular function.
Secondly, we have used these design guidelines to engineer materials to explore the importance of controlling the nano-spacing of vaccine adjuvants for selective activation of immune cells. Here, we coupled knowledge of pre-existing crystallography data of immune receptors to guide the design of spatially controlled ligand arrays on DNA-based nanomaterials. We created nanoparticles presenting CpG adjuvants to activate Toll-like Receptor 9 (TLR9) in a rationally designed way. Nanoscale variations in ligand spacing were shown to produce significantly different immunological responses: by matching CpG spacing to the distance of the TLR9 active dimer, cellular activation was significantly enhanced compared to random ligand presentation or free form CpG. For the first time, we show the tight connection between geometry and control of cellular signaling for CpG adjuvants. These findings are fundamental not only for a fine-tuned manipulation of the immune system, but also show a need to consider the importance of spatially controlled presentation and nanorigidity of therapeutics to increase efficacy and specificity of immune-modulating nanomaterials where multivalent binding interactions are involved.
We are currently exploring in more depth the potential avenues to use stabilizing coatings as transporter of our DNA nanomaterials into precise locations in the cytoplasm, as this is a rich pool of DNA-binding proteins. We aim to generate a toolbox that allows for an a-priori design of the best combination of coating / geometry / stability / uptake for a desired intracellular function.
Within the 3rd work package, exciting new collaborations are being created which have the potential to lift the research far beyond the state of art.
We are not in the position to disclose detailed speculations at this point in time.
We are not in the position to disclose detailed speculations at this point in time.