Using SSI Pools to Quickly Initiate Pandemic Molecule Manufacturing Campaigns for Phase I Clinical Trial
Site-specific integration (SSI) technology has brought forward much greater efficiency in inserting genes of interest (GOIs) into the host cell genome than the classical random integration transfection methods. While SSI has greatly improved the productivity of the stable cell lines, the much higher efficiency of recombinant gene integration into the genome of the host cells than the traditional random insertion has even greater pragmatic value for the industry. While the traditional method of plating and screening of stable cell pools can take up to 3 months from transfection to the fed-batch culture for a material generation, the SSI stable pool takes as short as 3 weeks for the same scale of production. This enabled us to conceive a more aggressive but pragmatic CMC timeline for counter-pandemic projects like neutralizing antibody productions. Since the transfections are performed under cGMP conditions, the SSI transfectants can be directly expanded for the production of toxicology materials and even one or two following batches of GMP production (Gen1 GMP), depending on the calculated needs for the initial clinical trials. From the perspective of CLD, which is ultimately intended for the creation of a high-quality, monoclonal production cell line, prioritizing the closest product quality proximity to the Gen1 GMP materials will be necessary for the top clone selection criteria, to control the product quality consistency between the Gen1 GMP batch materials and the following clonal GMP batch materials (Gen2 GMP). The MCB of the final clone (Gen2 GMP path) will eventually be used to succeed the Gen1 GMP production for the subsequent supply of clinical materials. The high-level flowchart for such a strategy is illustrated in Fig. 1.
Our SSI pools have proven to be quite capable of productivity on several toxicology or GMP productions with COVID-19 molecules (Fig. 2A). One representative toxicology production, which was directly expanded after transfection, yielded harvested titer as high as 4 g/L on day 14 with good growth and metabolic profiles, such as viability, viable cell density, glucose level, lactate level, and osmolality (Fig. 2B-F).
The comparability between toxicology and clinical materials is undoubtedly critical, one of the key challenges to overcome if the stable pools, instead of the monoclonal cell banks, should be used for manufacturing. The application of the SSI technology to such material generation strategy has an underlying prerequisite that the SSI pools should display stable product quality profiles, which we have observed with different seed train lengths for different batches of productions (data not shown). As illustrated in Figure 3, the toxicology batch and the following two GMP clinical batches of the same SSI pool (Molecule A) with different seed-train lengths shared similar productivity and product quality profiles to the smaller-scale bioreactor evaluations. In this case, the required amount of the appropriate materials was generated with satisfactory consistency, which also obviated the necessity for the purification and qualification protocol alterations.