Downstream Process Development for Adenovirus Vectors

With the increasing role of viral vectors and viral vaccines in modern medicine, the development of viral vectors represented by adenoviruses has become an important target for biomedical research. As the Food and Drug Administration (FDA) and the European Medicines Evaluation Agency (EDEA) continue to improve the requirements of drug safety, the development of viral vectors has become more complex, and developers need to seek the help of professional organizations. Creative Biolabs is a biotechnology company with leading technology in viral vectors preparation, and we meet your viral vector development requirements with strict quality control.

Overview of Adenovirus Vector Downstream Development

Since the first time that gene therapy products were approved for clinical trials in 1990, the viral vector technology as a gene delivery tool has developed rapidly, but there are still many places that need to be improved. Purification of well-functioning adenoviral vectors by rapid, simple and inexpensive downstream development processes has been our goal. How to mass-produce viral vectors that meet regulatory requirements for purity and biological activity is the first consideration in the downstream process of adenovirus development. In order to solve this problem, we need to have an in-depth understanding of the physical, chemical and biological characteristics of the viruses and cells utilized in the development process.

Figure 1. Downstream Bioprocess Development for Adenovirus Vector.

Adenovirus Vector Downstream Bioprocess Development Services

• Adenovirus capture step. Adenoviruses are not suitable for miniaturized high-throughput screening such as 96-well filter plates. We use the AKTA Pure 25 Chromatography System to perform dynamic binding capacity (DBC) screening on 1 ml pre-filled HiTrap^TM columns. The membrane was subjected to dynamic binding capacity (DBC) screening in a 1 ml ready-made membrane adsorber Q capsule. The concentrations of DBC in 20 mM Tris, ph 8.0 at different NaCl concentrations were determined. Tangential flow filtration (TFF) was added to the columnar solution with a residence time of 10 minutes and a 0.2-minute capsule fluid until the virus broke through.
• Optimization of elution conditions. The DBC screening results were further evaluated with adhesive resin or adsorbent to evaluate the step elution conditions of these adsorbents. After the loading step, a washing step with as high NaCl concentration as possible without virus penetration is required, followed by a step of complete virus elution with as low conductivity as possible to avoid co-elution of DNA with the virus.
• Resin evaluation for adenovirus polishing step. The materials purified in the evaluation capture step were studied using Capto Q, Capto Q ImpRes, and ready-to process adsorbent Q as samples. The sample was loaded at 0.1 CV and a 30 CV sample was loaded onto the Capto Core 700 using the agarose fast flow method. To determine the maximum sample size of Capto Core 700 with acceptable virus purity, fractions were collected and analyzed for impurity content throughout the sample loading phase.
• Scale-up of adenovirus capture and polishing steps. Divided into two processes, the first is to treat the sample with Capto Q ImpRes anion exchange resin and Capto Core 700 resin; an expanded cell culture harvest was then obtained using a combination of NFF sample and ReadyToProcess adsorber Q anion exchange membrane and Sepharose 4 fast flow size exclusion chromatography (SEC) resin.

Creative Biolabs combines a viable, rational downstream process route to design specific protocols based on customers' requirements. We will consider the design process from the overall production, economic and time cost, product purity, process stability, etc. If you have any questions or have any difficulties, you can contact us by email or send us an inquiry to find a complete solution.