Use of a viral mucin-like protein to convert adherent cells to suspension culture

DESCRIPTION (provided by applicant): Most cell culture lines are anchorage-dependent and require surface attachment for proliferation. For industrial production, increased surface area can be provided by microcarrier beads, but the associated increases incost and the resulting complexity of manipulation may preclude their use. Due to these limitations, anchorage-independent cells are preferred for the production of biopharmaceuticals, but appropriate anchorage-independent cells are not available for all applications; e.g., production of vaccines and cell-type specific proteins. This proposal explores the use of a novel insect virus protein to transform adherent cells to cells that can thrive in suspension culture. ParaTechs has identified a cell line fromAgrotis ipsilon (black cutworm), which provides levels of recombinant protein expression that are 3-to-10 times higher than the standard Sf9 cell line. Unfortunately, the cells are strongly adherent, which makes them unsuitable for large-scale protein production. We intend to stably transform the A. ipsilon cells with an insect virus gene that causes a loss of adhesion in hemocytes. We predict that expression of this protein in transformed cells will enable them to grow in suspension culture. In combinationwith our Vankyrin-Enhanced Baculovirus Expression Vector System (VE-BEVS), which has the ability to increase protein production per cell by a factor of 4 to 22, we anticipate achieving a level of protein expression per cell that is at least 12-to-220 times higher than currently possible. This dramatic increase in yield will be significant for all BEVS users, from individual researchers to large biopharmaceutical companies. We also will apply this technology to adherent mammalian tissue cultures. A simple method for converting mammalian cells from anchorage-independent to suspension culture would make a significant contribution to the production of vaccines and bio-therapeutics, and would result in improved human health. ParaTechs personnel are experiencedbaculovirologists and cell biologists. The protocols use standard technologies that are routinely adopted in our company. We do not anticipate difficulty with the experimentation. Current literature strongly supports our hypothesis and we are confident ofour success. PUBLIC HEALTH RELEVANCE: The future of human medicine will involve a dramatic increase in development of protein-based drugs and subunit vaccines, whose production will require large-scale propagation of tissue culture cells in suspension culture. Most continuous cell lines, however, are anchorage-dependent, and no method exists to routinely and easily transform adherent cells to suspension culture. ParaTechs will test an insect virus gene that causes hemocytes to lose adhesion for the ability to adapt anchorage- dependent insect and mammalian cells to suspension culture. Achievement of this goal would make a significant contribution to biopharmacology and human health.