Therapeutic proteins with posttranslational modifications, such as monoclonal antibodies (mAbs), are usually produced using clonally derived, stable mammalian cell lines. The generation of these cloned cell lines necessitates a time-consuming cell line development (CLD) process prior to the evaluation of the therapeutic protein's potential in clinical studies, consequently delaying patient access to novel therapies. This study aimed to accelerate and improve the CLD process. Initial efforts aimed at increasing the cell line productivity through the incorporation of a 1.5 kb ubiquitous chromatin opening element (UCOE) into the expression vector. This did so far not fully exhibit the intended effect, as stable chinese hamster ovary (CHO) cell pools containing the UCOE vector demonstrated lower productivity compared to control cell pools. However, at least the expansion phase was notably shorter for cell pools with UCOE vector, which might hint at a potential application in processes with strict time constraints. Furthermore, the CLD process was supposed to be accelerated through the application of a vector carrying a pac gene in conjunction with puromycin for selection instead of G418. This approach did not result in a shorter CLD process, however, yielded stable cell pools with a higher productivity compared to the control pools. Furthermore, the process performance of stable cell pools was compared to their cloned cell lines. Stable cell pools demonstrated competitive productivity relative to cloned cell lines in small-scale and fed-batch cultivation. The analysis of protein quality attributes by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), size-exclusion highperformance liquid chromatography (SE-HPLC), enzyme-linked immunosorbent assay (ELISA) and gel-based isoelectric focusing (IEF) demonstrated that a pool of stable CHO cells could produce mAbs with qualities comparable to those of cloned cell lines. This indicates that the application of a well-characterized stable pool for therapeutic protein production could be feasible to deliver protein in a quality sufficient for preclinical research and first-in-human (FIH) clinical trials. Simultaneously, single-cell cloning of this stable cell pool would be performed to generate cloned cell lines for manufacturing of qualitatively equivalent therapeutic protein for later clinical phases and beyond. It was demonstrated on shake flask scale that cloned cell lines can be generated with comparable cell culture parameters to that of the stable cell pools from which they derived. In summary, the CLD process was shortened utilizing a UCOE vector and the productivity was improved by using puromycin for selection in combination with a pac containing vector. Process performance data of stable cell pools compared to their cloned cell lines clearly indicated, that stable cell pools can produce mAbs with comparable quality parameters to their cloned cell lines.
Luis Schroer (Mon,) studied this question.