NK-cell Biofactory is an off-the-shelf allogeneic platform for cell-based synthesis of desired protein following recognition of a target cell while conserving the cytolytic activity. The molecular specificity of the platform can be redirected for treating a broad range of diseases.
The NK-92MI, a fast-growing cytolytic cell line with a track record of exerting clinical efficacy, is transformed into a vector for synthesizing calibrated amounts of desired engineered proteins at our disease site, that is, NK-cell Biofactory. This provides an allogeneic option to the previously published T-cell-based living vector that is limited by high manufacturing cost and product variability. The modularity of this pathway, which combines a “target” receptor with an “effector” function, enables reprogramming of the NK-cell Biofactory to target diseases with specific molecular biomarkers, such as cancer, viral infections, or auto-immune disorders, and overcome barriers that may affect the advancement of NK-cell therapies.
Parijat Bhatnagar
SRI Author
Lentivirus Manufacturing Process for Primary T‐Cell Biofactory Production
A process for maximizing the titer of lentivirus particles, deemed to be a necessity for transducing primary cells, is developed. Lentivirus particles, with a set of transgenes encoding an artificial cell‐signaling pathway, are used to transform primary T cells as vectors for calibrated synthesis of desired proteins in situ, that is, T‐cell biofactory cells. The process is also used to generate primary T cells expressing antigen‐specific chimeric antigen receptors, that is, CAR T cells. The two differently engineered primary T cells are expanded and validated for their respective functions, that is, calibrated synthesis of desired proteins upon engaging the target cells, which is specific for the T‐cell biofactory cells, and cytolysis of the target cells common to both types of cells. The process is compliant with current Good Manufacturing Practices and can be used to support the scale‐up for clinical translation.
Engineered Ovarian Cancer Cell Lines for Validation of CAR T Cell Function
A set of genetically engineered isogenic cell lines is developed to express either folate receptor alpha or mesothelin, and a control cell line negative for both antigens. These cell lines also express fluorescent and bioluminescent reporter transgenes. The cell lines are used to authenticate specificity and function of a T‐cell biofactory, a living vector that is developed to express proportionate amounts of engineered proteins upon engaging with disease cells through their specific antigenic biomarkers. The engineered cell lines are also used to assess the cytolytic function and specificity of primary T cells engineered with chimeric antigen receptors; and the specificity of monoclonal antibodies. The strategy described can be used to generate other cell lines to present different disease‐specific biomarkers for use as quality control tools.
Modular Antigen-Specific T-cell Biofactories for Calibrated In Vivo Synthesis of Engineered Proteins
An artificial cell-signaling pathway is developed that capitalizes on the T-cell’s innate extravasation ability and transforms it into a vector (T-cell Biofactory) for synthesizing calibrated amounts of engineered proteins in vivo. The modularity of this pathway enables reprogramming of the T-cell Biofactory to target biomarkers on different disease cells, e.g. cancer, viral infections, autoimmune disorders. It can be expected that the T-cell Biofactory leads to a “living drug” that extravasates to the disease sites, assesses the disease burden, synthesizes the calibrated amount of engineered therapeutic proteins upon stimulation by the diseased cells, and reduces targeting of normal cells.
