Periodic Reporting for period 1 - OBGate (Creating an orthogonal gate to the brain)
Periodo di rendicontazione: 2023-06-01 al 2025-11-30
The blood-brain barrier (BBB) is a tightly regulated interface that protects the brain but also severely limits the delivery of therapeutic molecules. Despite decades of research, most current strategies for crossing the BBB rely on hijacking endogenous transport systems, such as the transferrin receptor, which are already occupied by natural ligands and expressed in other tissues. This results in low specificity, competition, and potential off-target effects, making the safe and effective delivery of drugs and gene therapies to the brain an unsolved challenge in medicine.
The OBGate project ("Creating an Orthogonal Gate to the Brain") aims to revolutionize how therapeutics are delivered across the BBB. Our objective is to engineer a synthetic, orthogonal receptor expressed only in brain endothelial cells that enables the targeted transport of therapeutic cargo without interfering with the body’s natural systems. This “orthogonal brain gate” offers a programmable and specific mechanism to control what enters the brain, paving the way for more effective treatments of neurological diseases, including brain cancer and neurodegenerative disorders. In parallel, we are developing advanced lipid nanoparticle systems that can selectively deliver genetic material encoding the orthogonal receptor. These particles are designed to remain inert during circulation and activate only in specific brain environments, further enhancing targeting and safety. Together, these technologies will provide a new platform for overcoming the BBB and enabling a new generation of brain-targeted therapies.
The OBGate project ("Creating an Orthogonal Gate to the Brain") aims to revolutionize how therapeutics are delivered across the BBB. Our objective is to engineer a synthetic, orthogonal receptor expressed only in brain endothelial cells that enables the targeted transport of therapeutic cargo without interfering with the body’s natural systems. This “orthogonal brain gate” offers a programmable and specific mechanism to control what enters the brain, paving the way for more effective treatments of neurological diseases, including brain cancer and neurodegenerative disorders. In parallel, we are developing advanced lipid nanoparticle systems that can selectively deliver genetic material encoding the orthogonal receptor. These particles are designed to remain inert during circulation and activate only in specific brain environments, further enhancing targeting and safety. Together, these technologies will provide a new platform for overcoming the BBB and enabling a new generation of brain-targeted therapies.
In the first two years of the OBGate project, we have successfully created and validated OBGate 1.0 the first version of our synthetic receptor. Built on a modified natural receptor scaffold, OBGate 1.0 contains mutations that prevent it from binding to natural ligands while allowing it to recognize synthetic ligands specifically designed for therapeutic delivery. We demonstrated that OBGate 1.0 can be efficiently expressed in multiple human brain endothelial cell types and is functional in internalization assays.
We are also working on improved versions, which explore a variety of chimeric receptors to enhance receptor expression and trafficking. Advanced cellular assays and proteomic analyses are being used to better understand the intracellular pathways used by these receptors, and to identify protein interactors that could influence transport across the BBB.
On the delivery side, we shifted from polymer-based systems to more clinically relevant lipid nanoparticles and achieved efficient receptor expression using these carriers. A major innovation has been the development of conditionally activatable LNPs: these are coated with a peptide “mask” that prevents off-target effects and becomes activated only in the presence of tissue-specific stimuli. We also created new lipids that allow flexible attachment of targeting ligands, improving delivery specificity and versatility.
Together, these advances form the technical foundation for future in vivo testing and potential therapeutic applications.
We are also working on improved versions, which explore a variety of chimeric receptors to enhance receptor expression and trafficking. Advanced cellular assays and proteomic analyses are being used to better understand the intracellular pathways used by these receptors, and to identify protein interactors that could influence transport across the BBB.
On the delivery side, we shifted from polymer-based systems to more clinically relevant lipid nanoparticles and achieved efficient receptor expression using these carriers. A major innovation has been the development of conditionally activatable LNPs: these are coated with a peptide “mask” that prevents off-target effects and becomes activated only in the presence of tissue-specific stimuli. We also created new lipids that allow flexible attachment of targeting ligands, improving delivery specificity and versatility.
Together, these advances form the technical foundation for future in vivo testing and potential therapeutic applications.
Synthetic orthogonal receptor for BBB transport: We created the first synthetic receptor that is orthogonal while retaining the ability to be expressed and internalized by brain endothelial cells. This departs from traditional strategies that rely on hijacking natural receptors, offering greater selectivity and safety.
Activatable lipid nanoparticles: Our conditionally masked LNPs represent an innovation in gene delivery. These particles remain inactive during systemic circulation and only activate in specific environments (e.g. brain vasculature), greatly reducing the risk of unintended effects.
Modular ligand conjugation system: By synthesizing custom lipids with unique chemical handles, we introduced a flexible, site-specific method to decorate LNPs with targeting ligands. This allows for easy adaptation of the delivery system to different biological targets.
Efficient gene delivery in hard-to-transfect cells: We optimized nucleofection protocols to deliver DNA into endothelial cells with higher efficiency, enabling rapid and reproducible screening of receptor variants overcoming a common limitation in BBB research.
Establishment of robust platforms and tools: We built a suite of complementary tools for receptor design, cellular testing, and nanoparticle engineering. These can be applied beyond OBGate, supporting broader research into BBB transport and targeted delivery.
Activatable lipid nanoparticles: Our conditionally masked LNPs represent an innovation in gene delivery. These particles remain inactive during systemic circulation and only activate in specific environments (e.g. brain vasculature), greatly reducing the risk of unintended effects.
Modular ligand conjugation system: By synthesizing custom lipids with unique chemical handles, we introduced a flexible, site-specific method to decorate LNPs with targeting ligands. This allows for easy adaptation of the delivery system to different biological targets.
Efficient gene delivery in hard-to-transfect cells: We optimized nucleofection protocols to deliver DNA into endothelial cells with higher efficiency, enabling rapid and reproducible screening of receptor variants overcoming a common limitation in BBB research.
Establishment of robust platforms and tools: We built a suite of complementary tools for receptor design, cellular testing, and nanoparticle engineering. These can be applied beyond OBGate, supporting broader research into BBB transport and targeted delivery.