TWiP 244: Engineering Toxoplasma gondii for Intracellular Delivery of Therapeutic Proteins
0:00 Introduction: Introduces guest Lilach Sheiner, a professor at the University of Glasgow, who will discuss her research on using Toxoplasma gondii to deliver therapeutic proteins to neurons.
0:45 Toxoplasma gondii Life Cycle: Sheiner explains the life cycle of T. gondii, highlighting its ability to infect a wide range of warm-blooded animals, cross the blood-brain barrier, and establish dormant cysts in the brain.
7:44 Brain Invasion: Sheiner discusses the proposed mechanisms by which T. gondii invades the brain, including active invasion of cells and hijacking immune cells.
8:35 Engineering T. gondii for Protein Delivery: Sheiner outlines the challenges of delivering therapeutic proteins to the brain, particularly size and biochemical properties. T. gondii overcomes these challenges by producing proteins within the brain after crossing the blood-brain barrier.
15:20 T. gondii Secretion Systems: Sheiner details the two secretion systems of T. gondii that are exploited in this study: Rhoptry proteins (ROP) and dense granules (GRA). ROP proteins are injected into host cells, while GRA proteins are secreted after the parasite invades a cell.
16:29 Designing Fusion Proteins: Sheiner describes the process of designing fusion proteins that combine therapeutic proteins with either ROP or GRA proteins, allowing the parasite to secrete the therapeutic proteins into target cells.
20:22 Cell Culture Experiments: Initial experiments using fibroblasts (skin cells) in culture demonstrate successful expression and secretion of the fusion proteins, confirming the functionality of the system.
21:05 Dual Delivery System: Sheiner highlights the ability to utilize both ROP and GRA systems simultaneously to deliver multiple therapeutic proteins.
23:24 Neuronal Cell Experiments: The research team successfully delivered therapeutic proteins to primary neuronal cells in culture, demonstrating the system's potential for treating neurological disorders.
24:48 Brain Organoid Experiments: Experiments using brain organoids, which are 3D cultures that mimic brain structure, further validate the system's ability to deliver proteins to specific brain regions.
26:09 Mouse Experiments: In vivo experiments in mice demonstrate that the engineered T. gondii can deliver proteins to various brain regions, including the cortex, hippocampus, and brain stem.
29:27 Brain-Wide Delivery & Limitations: The research team used a Cre recombinase-based system to visualize the extent of T. gondii interactions with brain cells, revealing widespread interaction. However, the exact percentage of cells receiving the therapeutic protein remains to be determined.
31:04 Future Directions: Sheiner outlines future research directions, including:
Understanding and mitigating the virulence of T. gondii to ensure safety.
Optimizing the fusion proteins for more efficient delivery.
Exploring the use of less virulent T. gondii strains, such as the S48 strain used in sheep vaccines.
Investigating the factors that determine T. gondii’s cell and tissue tropism.
Conducting experiments in animal models with specific neurological defects to assess the therapeutic efficacy of the system.
39:28 Other Research in Sheiner’s Lab: Sheiner briefly describes her lab's research on the divergent mitochondrial biology of T. gondii and its implications for drug discovery.
46:22 Career Alternatives: Sheiner shares that if she were not a scientist, she would have considered a career in law, potentially as a judge, due to her interest in justice.