Understanding and manipulating lactate metabolism in single cells

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Many of the medicines prescribed today are proteins that are manufactured in animal cell cultures. Decades of research has examined ways to increase the productivity of these cultures by changing the nutrients the cells are fed, genetic engineering of the cells themselves and other strategies. However, there are still challenges that limit the overall amount of protein produced. One of these challenges is the accumulation of the metabolic waste product, lactic acid, the same waste product that causes your muscles to be sore after strenuous exercise. The concentration of lactate in animal cell cultures changes over time. Early on, it accumulates due to rapid metabolism, but later the cells use it to make energy and amino acids. This is called the lactate switch. Different batches of cells undergo the lactate switch at different times and the trigger for flipping the switch is not well understood. However, with respect to the production of protein medicines, the lactate switch is a positive trait associated with higher protein production.The proposed work seeks to create a set of tools for understanding and manipulating the lactate switch. The first goal is to create a non-invasive indicator that allows measurement of lactate concentration in individual cells. This tool would allow us to measure the extent of variation between cells, something that is currently not possible with standard techniques. We will use it to understand how different culture conditions affect lactate accumulation in the cells to try and identify how much variability there is in each scenario. Overall, this information can be used to choose manufacturing conditions with less difference between cells.The second goal is to develop tools that allow us to change the expression level of individual enzymes associated with lactate production and consumption. With these tools, we will be able to test hypotheses about which enzymes are associated with the lactate switch by controlling them with an external signal. Once the enzymes are identified, we could use the system to control when the lactate switch is flipped, making sure that all cells in the culture do switch from lactate production to consumption and making sure the switch flips at the best time to ensure high protein production. Finally, we will merge the two sets of tools to create cells that sense their own lactate concentration and when it gets too high, regulate their own lactate metabolism genes. This will create cells that do not make too much lactate, which should increase the amount of protein that they produce. The 'self-regulating cells' will also serve as a model for how to do this with other traits of interest in the future. This project could, therefore, change the way cells are developed for manufacturing purposes.