458840 Use of Real-Time Analytics and Apoptosis Assays for Enhanced Understanding of Cell Growth and Viability in a CHO-Based Process for Monoclonal Antibody Production

Tuesday, November 15, 2016: 12:30 PM
Continental 5 (Hilton San Francisco Union Square)
Yongqi Wu1, Nobel Vale2, Vivian Deng1, Mina Chaudhry1, Dimuthu A. Jayawickrama2, Timothy Stevens2 and Frank Ritacco1, (1)Process Development - Upstream, Bristol-Myers Squibb Company, Bloomsbury, NJ, (2)Biopharmaceutics Process Analytical Sciences, Bristol-Myers Squibb Company, New Brunswick, NJ

In a cell culture process for biopharmaceutical manufacturing, monitoring cell growth and viability is crucial for understanding cell behavior, as well as for determining cell culture duration and timing of harvest. Standard industry practice typically includes daily sampling of bioreactors for cell count and metabolite profiles. However, daily measurements are often too infrequent to detect rapid changes in metabolite concentrations and viability, and may not capture early indications of decline in cell health. We have evaluated the use of apoptosis assays as well as in-line monitoring techniques to gain a more complete understanding of cell growth, health and viability throughout the cell culture process.

In this study, we have applied markers of early, middle and late apoptosis to evaluate changes in cell growth, health and viability during a fed-batch process in 5L bioreactors. The markers that were monitored included 7-AAD, Caspases, and TUNEL assay for DNA nicking. Multiple early and mid-stage apoptotic markers were compared for their effectiveness in evaluating cell health and predicting cell growth trends in a bioreactor. Additionally, to provide more frequent cell growth measurements and metabolite readings to align with apoptotic assays, in-line probes were incorporated for real-time data analysis. Through calibration and method development, a capacitance-based Incyte probe was used for real-time monitoring of viable cell density (VCD), and an in-line Raman spectroscopy probe was used for real-time monitoring of glucose and lactate.

The results demonstrated that apoptotic markers can provide additional insight into cell health beyond what is learned from daily cell counts, thereby greatly deepening our understanding of cell culture conditions and process parameters during development. Markers of early apoptosis were observed several days prior to a drop in viability as measured by a commercial cell counter, while late-stage apoptotic markers were observed hours in advance or concurrently with the VCD and viability drop. Further insight into the health and productivity of the culture may be obtained by correlation of in-line metabolite and cell growth profiles with cell apoptotic progress.

In summary, optimization and application of real-time analytics and apoptosis assays in a CHO cell process can provide a rich and complete data set for better visualization of cell growth and health throughout the culture process, thereby enabling process improvement to maximize product titer and quality.

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