End-to-end process support, from development to industrialization
Lentiviral vectors are one of the most commonly used viral vectors for therapeutic purposes due to their specific functional properties, such as their ability to infect non-dividing cells and packaging capacity. The RNA-based vector has been designed to deliver genes to mammalian cells safely and effectively, and there are several therapies using lentiviral vectors currently under development and in human clinical trials. Developing an efficient process for producing a lentiviral vector-based therapy is not easy. Purification of these 80-120 nm envelope vectors is challenging due to their low stability.
There are multiple strategies for developing lentiviral vectors, but most follow the same basic process, with some variation as needed.
Adherent Cell Seeding
Early on in therapeutic development, viral vectors are usually manufactured in adherent cells using two-dimensional systems such as cell factories or roller bottles. As processes scale up and more virus is needed, the footprint and manual operations required to grow cells in this fashion quickly becomes problematic.
The Xpansion Multiplate Bioreactor is a closed, single-use system that has been specifically designed for the scale-up of shear-sensitive, adherent mammalian cells under controlled conditions. It provides a similar microenvironment to a cell factory but in a much more compact space and with fully automated environmental control.
Adherent Cell Culture
As the therapies progress into clinical development, there is a requirement to increase quantity and quality of the virus produced. Manufacturing virus from adherent cells using fixed-bed bioreactor technology allows for direct transfer of the 2D reference process, which minimizes risk and saves time.
The iCELLis bioreactor is a fully-integrated, high-cell density bioreactor designed to simplify processes involving adherent cells by combining the advantages of single-use technologies with the benefits of a bioreactors - reduced handling, footprint, labor, cost, and improved process assurance, robustness, and scalability. The iCELLis technology can be used at bench-scale (0.5 to 4 m2) and manufacturing-scale (66 to 500 m2).
Suspension Cell Culture
Manufacturers are increasingly looking to produce viral vectors for gene therapy in suspension cell cultures, as the pressure increases for increased scalability. Viral vector production in suspension culture usually requires additional testing and optimization. Our experienced team of applications experts can work with you to determine the best solution for your upstream processes.
Allegro™ STR Bioreactors
Allegro STR single-use stirred tank bioreactors were thoughtfully designed to maximize usability and process assurance while delivering consistent and scalable cell culture performance across a full range of maximum working volumes, from 50 – 2000 L.
Allegro™ XRS Bioreactors
The Allegro XRS 25 bioreactor system is a single-use bioreactor system with unique agitation and control properties designed for the cultivation of mammalian cells in suspension culture under controlled conditions at the 2 to 25 L scale.
The clarification step removes cells, cell debris, and other impurities to reduce biological burden. The easiest and most economical technology to clarify the cell culture is filtration. Low yields are a common challenge in this step, especially as cell densities increase upstream. Our experienced technical team can help you find the best filter technologies and protocols for your process, whether you are just beginning process development or need to re-evaluate and optimize this step down the line.
Supor® and Fluorodyne® membranes are appopriate for this step and are available in a wide range of sizes and formats. Contact us for assistance in determining the best filter technology for your process.
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Ion exchange purification is the next step in this downstream process, to remove empty capsids, DNA and host cell proteins (HCP). Adsorptive membrane technology offers an efficient and effective option for this step.
Mustang® Q Membrane Chromatography Capsules
Ultrafiltration/Diafiltration using Tangential Flow Filtration (TFF) membranes further concentrates the target molecule and exchanges buffers. Achieving high yield is a common challenge with TFF, but our technical experts can work with you to optimize your UF/DF unit operation based on experience and thorough testing.
Single-Use Tangential Flow Filtration (TFF) Modules
Single-use tangential flow filtration (SUTFF) modules are ready-to-use gamma-irradiated cassette modules that can be integrated into single-use TFF set-ups. Integrating the process-proven Omega™ polyethersulfone (PES) membrane, single-use TFF modules are available in scalable formats from 93 cm² up to 2.5 m² of membrane area.
T-Series TFF Cassettes with Omega Membrane
T-Series cassettes with Omega membrane are ideal for development, pilot, and production-scale TFF applications in diverse biological and biopharmaceutical processes. They are especially useful in vaccine and conjugate concentration and diafiltration, purification and recovery of monoclonal antibodies (mAb) or recombinant proteins, and blood plasma fractionation and purification.
T-Series TFF Cassette Holders
TFF cassette holders for the medium and large-scale production environment have a small footprint, yet still are capable of holding large areas of membrane. All stainless steel holders are engineered to precise specifications with a high standard internal and external surface finish and are available with either manual torque (MT) or auto torque (AT) compression systems for ease of use.
Bulk Drug Substance Filtration
Manufacturing for clinical purposes requires the production of highly pure and biologically active vectors that meet regulatory requirements. A last sterilizing grade (0.2 µm) filtration step is usually needed before product is sent for final filling.
Sterile Liquid Filtration
Supor® EKV, Supor EX ECV and Fluorodyne® EX EDF membranes are appropriate for this step and are available in a wide range of sizes and formats. Contact us for assistance in determining the best filter technology for your process.
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Pall has a global team of over 300 experienced technical professionals to support your molecule from pre-clinical to commercial manufacturing. From process development services and end-to-end platforms, to on-site testing and applications support for specific process steps, Pall will help you find the best solution based on data and testing.