Labisch Jennifer J, Evangelopoulou Maria, Schleuß Tobias, Pickl Andreas
Lab Essentials Applications Development Sartorius Göttingen Germany.
Institute of Technical Chemistry Leibniz University Hannover Hannover Germany.
Eng Life Sci. 2025 Jan 3;25(1):e202400057. doi: 10.1002/elsc.202400057. eCollection 2025 Jan.
The demand for lentiviral vectors (LVs) as tools for ex vivo gene therapies is ever-increasing. Despite their promising applications, challenges in LV production remain largely due to the fragile envelope, which challenges the maintenance of vector stability. Thus, downstream processing optimization to enhance efficiency, yield, and product quality is necessary. This study investigated the influence of membrane types and filtration devices during ultrafiltration (UF). Nine different membrane materials consisting of polyethersulfone (PES), regenerated cellulose, or Hydrosart, with distinct molecular weight cutoffs, were evaluated in stirred cells, centrifugal ultrafilters, and crossflow cassettes. The evaluation was based on the ability to retain infectious LV particles and remove impurities. The analysis revealed that a reinforced 100 kDa PES and a 300 kDa Hydrosart membrane had the best overall ability to concentrate infectious LVs and remove DNA, especially when operated in a stirred cell. Challenges were seen in the nonoptimized crossflow cassette process, where infectious LV recovery was generally lower compared to other devices. We demonstrated that membrane material and filtration device have a direct impact on the efficiency of LV UF.
作为体外基因治疗工具的慢病毒载体（LV）的需求一直在增加。尽管其应用前景广阔，但LV生产中的挑战仍然主要源于脆弱的包膜，这对载体稳定性的维持构成了挑战。因此，有必要进行下游加工优化以提高效率、产量和产品质量。本研究调查了超滤（UF）过程中膜类型和过滤装置的影响。在搅拌式细胞、离心超滤器和错流盒中评估了九种不同的膜材料，包括聚醚砜（PES）、再生纤维素或Hydrosart，它们具有不同的截留分子量。评估基于保留感染性LV颗粒和去除杂质的能力。分析表明，增强型100 kDa PES膜和300 kDa Hydrosart膜在浓缩感染性LV和去除DNA方面具有最佳的整体能力，尤其是在搅拌式细胞中操作时。在未优化的错流盒工艺中发现了挑战，与其他装置相比，感染性LV的回收率通常较低。我们证明了膜材料和过滤装置对LV超滤效率有直接影响。
