SBIR Phase I: Novel Compact Cell Settler for Perfusion Cultures of Microbial Cells

这一小型企业创新研究(SBIR)项目的更广泛影响/商业潜力是使用更高效的生产技术降低治疗性生物制剂的制造成本。这将通过开发新的细胞保留技术来实现，以提高重组微生物和哺乳动物细胞的连续灌流生物反应器培养中的细胞浓度和生产率。基于已经在大规模哺乳动物细胞培养中被证明的倾斜沉淀器技术，正在开发紧凑型和更高效可扩展的细胞或颗粒沉淀器，以将其应用扩展到其他工业领域，如微生物酵母分泌生产系统、多相催化、城市净水、废水处理等。在此阶段I拨款中，一种用于重组微生物酵母细胞的新型紧凑型细胞沉淀器设备得到了彻底的表征，它将被推出用于大规模生产重组微生物和哺乳动物细胞的生物制品。随着生物制造技术从早期的补料分批培养成熟到连续生产，由于市场规模的扩大和更一致的产品设计质量考虑，这些新型紧凑型澄清器将能够在大规模高细胞密度灌流生物反应器中更有效地生产许多治疗性生物制剂，并有助于显著降低从重组哺乳动物和微生物细胞培养的生产成本。这项SBIR第一阶段项目建议开发一种新的方法来扩大已证实的倾斜澄清器技术的规模，该技术在过去作为片状澄清器已被直线放大。虽然层状沉淀物已经成功地用于在高密度灌流生物反应器中生产重组哺乳动物细胞分泌的治疗性生物制品，但它们还没有被用于微生物细胞培养。这项建议的研究目标是开发更紧凑和高效的重组酵母细胞沉淀物，并表征这些沉淀物在收获过程中选择性地去除死亡细胞并将活的和生产的细胞回收回生物反应器的能力。第一个原型的初步结果显示，较小的酵母细胞被选择性地移除，而大多数较大的细胞被浓缩并回收回生物反应器。与生物反应器相比，这些紧凑型沉淀器在收获流中留下的酵母细胞浓度更低，尺寸分布更小，在稳定的连续培养中将测量到，由于活细胞和生产细胞完全循环到生物反应器，灌流生物反应器中的细胞密度将显著增加，这将在不同的灌流收获速率下得到表征。

The broader impact/commercial potential of this Small Business Innovative Research (SBIR) project is to reduce the manufacturing cost of therapeutic biologics using more efficient production technologies. This will be accomplished by developing novel cell retention technologies for increasing cell concentration and productivity in continuous perfusion bioreactor cultures of recombinant microbial and mammalian cells. Based on the inclined settler technology already proven in large scale mammalian cell cultures, compact and more efficiently scalable cell or particle settlers are being developed to expand its applications to other industrial areas, such as microbial yeast secretory production systems, heterogeneous catalysis, municipal water clarification, wastewater treatment, etc. Following thorough characterization of a novel compact cell settler device for recombinant microbial yeast cells during this Phase I grant, it will be launched commercially for large-scale manufacturing of biologics from recombinant microbial and mammalian cells. As biological manufacturing technology matures from early fed-batch cultures to continuous manufacturing due to increasing market size and more consistent product quality-by-design considerations, these novel compact settlers will enable more efficient production of many therapeutic biologics in large scale high cell density perfusion bioreactors and contribute to a significant reduction in cost of their manufacturing from recombinant mammalian and microbial cell cultures.This SBIR Phase I project proposes to develop a novel method of scaling up the proven inclined settler technology, which has been scaled up rectilinearly in the past as lamellar settlers. While the lamellar settlers have been used successfully to manufacture therapeutic biologics secreted by recombinant mammalian cells in high cell density perfusion bioreactors, they have not yet been adapted for microbial cell cultures. The research objectives of this proposal are to develop more compact and efficient settlers for recombinant yeast cells and characterize the capability of these settlers to remove dead cells selectively in the harvest and recycle the live and productive cells back to the bioreactors. Initial results from the first prototype demonstrate selective removal of the smaller yeast cells, while most of the larger cells are concentrated and recycled back to the bioreactor. Lower cell concentrations and smaller size distributions of the yeast cells leaving in the harvest stream from these compact settlers compared to the those in bioreactors will be measured in steady state continuous cultures and dramatically increasing cell density in the perfusion bioreactor due to the complete recycle of live and productive cells to bioreactor will be characterized for different perfusion harvest rates.