Role of autophagy in bioprocessing

Chinese Hamster Ovary (CHO) cells are the cells most frequently used in biomanufacturing for the production of biologics.   During a typical bioprocess the process terminates due to cell death which results in low levels of productivity.  Although many strategies have been developed to extend cell viability (life), detailed understanding of the mechanisms underpinning cell behaviour during the late stages of culture is still lacking.  Better understanding of cell physiology during the final stages of culture would be invaluable for identifying novel cell engineering strategies and optimal bioprocess conditions to extend cell viability, and thus, achieve higher protein yields.

Cell death through autophagy and apoptosis are commonly observed to occur simultaneously and experimental evidence for mammalian systems suggests that, when exposed to stress, the cells first engage autophagy as a survival response, and if the same stress is prolonged, apoptosis is triggered.  Our overall objective is to develop an understanding of how and why cells die during bioprocess for production of biopharmaceuticals.  Placenta-specific 8 (PLAC8), a cysteine-rich 12.5 kDa protein has been shown to have multiple and diverse roles in increasing or decreasing proliferative rate, resistance to apoptosis and autophagic activity depending on the tumour type.  This project examines the role of autophagy in bioprocessing using CHOK1 stably transfected with Plac8 and CHO-DP12 cells which produce an antibody.  Cells are grown in batch and fed-batch culture and cell productivity measure by ELISA and cell death (both apoptosis and autophagy) measured using flow cytometry (CYTO-1D and TO-PRO-3), western blot analysis for autophagy specific markers (e.g., p62, LC3-I, LC3-II) and confocal microscopy (Plac-8, Lamp1).  The effect of pharmacological inducers and inhibitors of autophagy on cell growth and productivity are also being examined as a potential strategy to overcome autophagy in bioprocessing.

Project Funding: UCD Advance PhD Core Scheme (2019-23)

PhD Researcher: Raymon Floyd Principe

PhD Supervisor: Susan McDonnell