The first phase of process development involved the high-throughput batch screening of chromatographic resins in bind-and-elute mode in a 96-well slurry plate format to identify the best performing candidates. The stability study helped set the operating condition boundaries of pH and salt for the subsequent evaluation of chromatographic resins. The selection of buffer species also impacted vector stability, with LVV exhibiting increased stability in phosphate buffer in the pH range of 6.5-7.5. While the vector was observed to be stable at pH 6.5-7.5, high salt concentration (> 500 mM NaCl) was found to be detrimental to vector stability. To address this, we first performed a detailed study to assess the stability of LVV under different fluid phase conditions and identified optimal conditions for vector stability. However, the purification process suffers from low yield mainly due to the poor stability of lentiviral vectors and low product titer in the feed material, resulting in limited scalability and high manufacturing cost. Lentiviral vectors are mainly used as gene therapy agents in cancer immunotherapy and for the treatment of neurodegenerative diseases. Later, a novel separation process was also designed for the isolation of cells for cell therapy applications. With this in mind, chromatographic purification process was developed for lentiviral and adeno-associated viral vectors, which are the two major vectors currently used in gene therapy-based clinical trials. As various gene and cell therapy-based treatments progress from pre-clinical to clinical and finally to the commercialization phase, the high demand for these new biologics necessitates developing robust, fast, and scalable downstream purification techniques that will guarantee optimum product yield, purity and viability. AbstractWith the recent progress in gene and cell-based regenerative therapies, the challenge of producing pure and concentrated stocks of complex biologicals, in particular, gene therapy vectors and functional cells, has gained significant importance.
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