Viral vectors for cell and gene therapy

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Viral vectors for cell and gene therapy

At VVector Bio we have years of experience and extensive expertise in the production, purification, and characterization of viral vectors for cell and gene therapy, viral based-vaccines, and recombinant proteins. Aside from our expertise in process development we also focus on delivering your final product in high quantities and quality. We can deliver these products for both in-vitro and in-vivo experiments to accelerate the path to the clinic of the project

 

Adeno-Associated Virus (AAV)

VVector Bio has extensive experience in the manufacturing of AAV by triple transient transfection in HEK-293 suspension cells.

 

We have the plasmids for Rep/Cap for different serotypes including Rep2/Cap 2, 5, 6, 8, and 9 and the plasmid containing the adenovirus helper function. You can start your project with just the gene of interest (GOI) sequence in hand and we will provide you with the final genetic  constructions, develop the production process, purification and perform the characterization and analytical quantification of the final product.

 

Furthermore, we have experience in house with cell assays to assess the functionality of the viral vectors using internal standards, so we can develop and validate your cell-based assay to determine the functional particles of your specific product.

 

VVector Bio also offers in-house technology for high-yield AAV production using the Baculovirus Insect cell system.  We provide the molecular construction of the Bacmid containing the GOI and amplification of baculovirus stocks Bac-Rep, Bac-Cap and Bac-GOI followed by co-infection in SF9 cells, purification and characterization of the final product.

Process package:

 

Production:

  • Triple transient transfection: (pGOI-ITR, pRepCap, helper plasmid) in HEK293 cells
  • 1L, 3L, 10L Bioreactors (1013 vgs/L in supernantent)

 

Purification:

  • Affinity chromatography
  • Anion exchange chromatography for enrichment of full particles.
  • Separation of full/empty particles is also available

 

► Analytical:

  • ddPCR
  • Western Blot
  • Electronic Microscopy
  • Total protein determination
  • NSEM
  • Host cell protein, Host cell DNA (on request)
  • Endotoxin level (on request)

Lentivirus

VVector Bio provides services to produce lentiviral vectors in HEK-293 suspension cell culture.

 

The lentivirus packaging is mediated by transient transfection of 4 plasmids for 3rd generation lentivirus and 3 plasmids for 2nd generation LV depending on your needs. The main difference between 2nd and 3rd generation lentivirus packaging systems is that the Rev element is in a separated plasmid for the 3rd generation lentivirus which makes them safer for the clinic.


At VVector Bio we implement a perfusion process intensification strategy to increase lentivirus vector yields and maintain stability. The cell culture supernatant is concentrated by a robust tangential flow filtration (TFF) process followed by anion exchange chromatography and sterile filtration. We also perform direct concentration by ultracentrifugation through sucrose cushion. The basic analytical characterization provided in the certificate of analysis for lentivirus consist of ddPCR, RCL, total protein concentration and host cell DNA.

Process package:

 

Production:

  • Quadruple transient transfection in HEK-293
  • Scale 1L, 3L, 10L bioreactor

 

Purification:

  • TFF
  • Anion exchange chromatography
  • Alternative concentration step by ultracentrifugation

 

► Analytical:

  • Quantification of virus particles  by ddPCR
  • p24 quantification by ELISA
  • Total protein
  • Host cell protein, Host cell DNA (on request)
  • Replication competent lentivirus (on request)
  • Endotoxin level (on request)

VVECTOR BIO, Bridging Research and Clinic