10086682Engineering of nature-mimetic sub-micron crystalline protein depots to incorporate high cargo densitiesClosedCollaborative R&DInnovate UKPurified recombinant proteins are widely used in research, medicine, and industry. Cytokines, hormones and antibodies have been important in reshaping the field of medicine over the last half-century. How these proteins are manufactured, stored and delivered to their site of action (e.g. diseased tissue) is fundamental to their utility. We have developed PODS, a nature-mimetic technology, that adds value to all aspects of a recombinant protein's life cycle. PODS are sub-micron scale cubic protein co-crystals (containing a cargo protein) produced in insect cells. PODS utilizes an innovative manufacturing technology, based on a process that naturally occurs in silkworms, to produce bioactive proteins incorporated into sub-micron scale protective crystals (made of a bio-inert polyhedrin protein). The crystals greatly simplify the purification of the recombinant protein, as they are physically distinct from other components of the production cell. Storage instability is addressed as cargo proteins are stabilized within the crystal lattice and have high levels of stability over many months, even at elevated temperatures in aqueous suspension. Finally, following administration, at the site of action, the crystals are degraded by proteases providing a matrix-degradation dissolution-based cargo release system. This provides sustained release of the soluble cargo protein over a one-month period. We have developed PODS for research and therapeutic applications, primarily for localized sustained release of cytokines. Most recently, we have demonstrated that PODS are able to harness phagocytic immune cells, such as monocytes, neutrophils and macrophages, to target the delivery of immunostimulatory cytokine proteins to cancer. We plan to extend this delivery approach to other protein classes, particularly peptides and small antibodies, known as nanobodies. However, the packaging efficiency of cargo proteins into PODS, constituting around 1% of the total protein, is a limitation: Whilst this is sufficient for delivering highly potent proteins such as cytokines, other protein classes, such as antibodies and anti-microbial peptides, require higher loading efficiency. In addition to therapeutic applications, there are industrial applications of PODS, such as the biomanufacturing of cultured meat, in which cost reductions enabled by increased cargo density would be highly advantageous. The project will focus on the bioengineering of PODS to generate a novel PODS architecture with increased cargo density. Two independent strategies will be assessed to create PODS that have a higher cargo protein loading capacity. These PODS will be tested in a range of assays to assess cargo packing density, release profile and bioactivity.