Valinhas, Ana Teresa de Sousa; (2019) Microscale processing tools as screening strategies for cell therapy manufacture. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

Allogeneic cell therapies require the manufacture of large amounts of cells in a reliable and robust manner. Microcarrier suspension cultures are scalable, providing a solution for this challenge. Screening the effects of raw materials on cell quantity and quality should be performed at early stages of process development to reduce the risks associated with scale-up. A microfuidic platform was investigated to measure quantity and phenotype of cells grown on microcarriers. This approach was tested using CTX0E03 cells and PA5 hOMCs, potential cell therapy candidates for the treatment of neurological diseases. In well plates Plastic L and Synthemax II LC microcarriers yielded 1.2 -1.5x10⁴ cells/cm². Expression of ß-III tubulin and nestin was maintained similar between the two conditions. For PA5 hOMCs, Plastic led to 3 times higher cell density compared to Plastic Plus microcarriers. The expression of b-III tubulin and nestin was 1.5 to 2-fold higher for both Plastic and Plastic Plus microcarriers compared to other tested conditions. In the second generation microfuidic platform, cell number and phenotype were investigated through an immunoassay and a tailored image processing tool. Results were comparable between platforms for cell quantity. CTX0E03 cells proliferated similarly on Plastic L and Synthemax II LC with cell densities ranging from 1.3-1.5x10⁴ cells/cm². For PA5 hOMCs, in the second generation microfuidic platform, the cell density on Plastic was 1.2 times higher compared to Plastic Plus. A study performed in stirred microcarrier cell culture showed 3 times higher cell density on Plastic microcarriers compared to Plastic Plus microcarriers. Furthermore, cells grown on Plastic Plus microcarriers led to 2-fold higher expression of p75NTR and signi cantly increased neurite outgrowth in a co-culture assay with NG108-15 neurons with 1.2 times higher average and maximum neurite length, and 1.7 times higher number of neurites per neuron.

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