Engineering the cell-free production of viral proteins
by Lauren Mamer
Can you envision a world in which cancer patients receive personalized vaccines carried by synthetically-produced viruses? With a new method of synthesizing viral proteins, Professor James Swartz in the Departments of Chemical Engineering and Bioengineering at Stanford is working to make this a possibility.
Engineering Viruses
Viruses encase their genetic material in protein shells called capsids that invade our cells. This infectious agent, however, can be used for our advantage. Swartz and graduate student Brad Bundy have recently engineered the production of viral proteins that assemble to form empty capsids. These virus-like particles lack the pathological effects of naturally occuring viruses and can potentially deliver targeted treatment to diseased cells.
Cell-Free Production of Capsids
Swartz's research group uses a cell-free system for the production of these viral proteins. Traditionally, proteins have been synthesized in whole cells, which must be kept alive in order to produce the proteins. Therefore, one disadvantage of using intact cells is that a large portion of metabolic resources is dedicated to sustaining the cell. In a cell-free environment, however, all raw materials and resources can be devoted to producing the desired protein. Furthermore, the protein can be produced at higher concentrations than in whole cells, where such high levels of protein are toxic to the cell.
To create an efficient cell-free system, Swartz and Bundy rip apart E. coli cells to harvest useful bacterial components, such as ribosomes and membrane vesicles that act as respiration centers to fuel protein production. They then use an RNA sequence coding for a specific viral protein, and optimize protein yield by inhibiting competing reactions and preventing the degradation of amino acids (building blocks of protein).
Besides providing high protein yields, the dilute cell-free system also facilitates proper protein folding by providing plenty of space and time for assembly. To assemble a capsid, 180 copies of the viral protein join together to form a Òsoccer ballÓ sphere that is 27 nm in diameter. Bundy has found that around 75% of the protein units produced in cell-free synthesis successfully assemble into capsids. Capsids as Vaccine Vehicles
These viral capsids can be used to transport vaccines that are attached to their surfaces. Aaron Goerke, a graduate student in Swartz's group, has chemically linked an unnatural amino acid that is uniquely reactive, such as azidophenylalanine, to the capsid. Via this amino acid, the capsid can react at biologically active conditions with an alkyne (triple-bonded carbon) on an immunogenic protein. Once the protein binds to the capsid, it can circulate throughout the body and will interact only with cells that have a corresponding receptor for this protein, thus producing a strong immune response that is specific to this protein.
Such targeted vaccines, coupled with the advantages of cell-free protein synthesis, could be especially helpful in the treatment of B cell lymphoma. This type of lymphoma affects the antibody-producing B cells. Each patientÕs cancer is different because there are many variable regions present on the receptors of the B cells. With Swartz's cell-free system, proteins that would induce protective responses against a patientÕs specific lymphoma could be tailor-made within two weeks, rather than the six months required using current synthesis methods. These immunogenic proteins could be attached to the viral capsids, which would then deliver the proteins directly to the cancerous B cells and elicit a faster, more specific immune response. Additionally, the engineered capsids avoid the side effects of traditional, generalized treatments for cancer, e.g., radiation and chemotherapy regimens.
Future Targets
The combination of cell-free synthesis and viral capsid packaging holds much promise for future medical applications. Swartz and his team developed this process with the goal of producing proteins on a commercial scale. Although the capsids have yet to withstand years of development and clinical testing before becoming a useful therapy, Swartz is Òvery excited about the potential versatility of this vaccine platform and believes that it will allow us to develop novel vaccines to elicit rapid and precise protective responses.Ó