A Scalable Lentiviral Vector Production and Purification Method Using Mustang Q Chromatography and Tangential Flow...

Area of Science:

• Biotechnology
• Molecular Biology
• Gene Therapy

Background:

• Lentiviral vectors (LV) are crucial for stable gene transfer due to their ability to transduce both dividing and quiescent cells.
• LV offer flexibility in cell-specific expression and tropism modification, enabling precise genomic manipulation.
• Current production often relies on adherent 293T cells and transient transfection, with subsequent purification steps.

Purpose of the Study:

• To describe a scalable pilot-scale manufacturing method for lentiviral vectors.
• To optimize the purification and concentration of lentiviral vector supernatant.
• To achieve high titers of infectious lentiviral particles for research and clinical applications.

Main Methods:

• Utilized adherent 293T cells for lentiviral vector production via transient transfection.
• Collected un-concentrated vector supernatant 24-48 hours post-transfection.
• Employed Mustang Q chromatography for purification, Tangential Flow Filtration for concentration, and diafiltration for final formulation.

Main Results:

• Successfully purified and concentrated approximately 6 L of supernatant to 150 mL.
• Achieved typical titers ranging from 1 x 10^8 to 1 x 10^9 infectious particles/mL.
• Demonstrated the method's scalability for increased volume and yield.

Conclusions:

• The described pilot-scale method provides an efficient approach for lentiviral vector manufacturing.
• This process yields high-titer lentiviral vectors suitable for gene therapy research.
• The method is amenable to iterative performance and further scale-up for larger production needs.