As the gene and cell therapy landscape has expanded there has been a subsequent expansion in the need large-scale production of GMP-grade AAVs that meet regulatory filing standards. PackGene aims to meet this need across phases of the gene and cell therapy lifecycle including PI initiated clinical trials (IIT), to Investigational New Drug (IND) filings, to Phase I-III clinical trials, and large-scale commercial production of GMP-grade AAV.
Using our proprietary platforms and technologies PackGene provides mass-production services for GMP-compliant AAV, as well as GMP production line set-up, virus processing and assays, and IND pharmacological documentation for processes ranging from the upstream rAAV generation to the downstream isolation and purification. We also ensure that our AAV manufacturing services comply with GMPs and all applicable regulatory and required standards for IIT and IND applications.
Note
PackGene’s FDA and EMA compliant AAV GMP manufacturing facility (~2000m²) begin production in 2021. For more information, please contact us.
PackGene GMP-compliant services include AAV upstream production as well as downstream development, purification, large-scale fermentation, material segregation, traceability record keeping, document control, and application of a comprehensive range of quality control assays.
PackGene has developed core processing technology that multiplies single-batch AAV output/yield and reduce input cost, thus giving our customers a leading edge.
AAV Quality Control (or customized based on specific process)
| Specification | Assay Methods | |
| Identity | GOI Sequence | Sanger Sequence |
| AAV Capsid | SDS PAGE/MS | |
|
Purity
|
HPLC Purity | HPLC |
| UV Purity | A260/A280 | |
| %Empty Capsids | AEX HPLC | |
| TEM | ||
| AUC | ||
| Aggregation | DLS | |
| Potency & Content | Capsid Titer | ELISA |
| Genome Titer | ddPCR | |
| Infectious Titer | TCID50 | |
| Protein Expression | In Vitro assay | |
| Impurity | Residual Host Cell DNA | qPCR |
| Residual E1A | qPCR | |
| HCP Residue | ELISA | |
| BSA Residue | ELISA | |
| Residual Plasmid | ddPCR | |
| Residual Nuclease | ELISA | |
| Residual Iodixanol/CsCl | HPLC | |
| Residual Ligand | ELISA | |
| Safety | Mycoplasma | qPCR/Mycoplasma culture /DNA staining |
| rcAAV | Cell Assay+qPCR | |
| Bacterial endotoxin | LAL | |
| Sterility | Direct Inoculation Method/Membrane filtration | |
| Adventitious agent | Cell Culture | |
| Abnormal toxicity | Animal test | |
| General Characterization | Appearance | Visual Inspection |
| Content | Minimum Fill | |
| pH | pH Method | |
| Osmolality | Osmometers | |
| Particulate matter | Light Obscuration Particle Count test |
Our GMP manufacturing site fully adopts “disposable technology” and advanced design concepts. Our facility is equipped with a clean environment in line with quality by design (QbD) concepts. This includes physical isolation and full-exclusion systems to segregate different functional workshops. The GMP workshop is only used for pilot testing or large-scale production of AAV vector, and this completely avoids multi-species sharing or cross-contamination issues commonly found in traditional CDMO platforms.
The upstream production line of our GMP site is equipped with the leading edge iCellis 500 disposable bioreactor system (up to 500m2 wall area), STR200T 200L disposable reactor, Xpansion disposable reactor, XDR 50 MO 50L disposable reactor, and several wave-shaped bioreactors to meet the needs of large-scale wall and suspension culture production of AAV. Our downstream line is equipped with ÄKTA chromatographic purification and ultrafiltration systems. Based on years of experience in large-scale production, single batch AAV yields of 1E+15-17GC or vg can be achieved, allowing for a shortened pilot production cycle of AAV under GMP standards that can be completed within 1-2 months. PackGene’s core process technology route can help partners to significantly increase single batch yields and reduce input costs. This results in valuable speed advantages for customers and ultimately leads to accelerated registration filings and clinical level production of gene therapy services.
The purpose of final generation cell assays is to confirm cell suitability and to determine if cells have mutated or produce carcinogenic substances. Cells are resuscitated and passaged from the working cell bank according to the production process route, transferred to the bioreactor for passaged digestion, cultured according to the proposed process, and then digested for lyophilization or sent for testing at the end of bioreactor production. The generation of cell resuscitation and passaging methods are carried out according to your custom production process SOP.
PackGene facility includes iCellis500 (~500m2 adhesion area), STR50/200/500T with the single-use technology that can flexibly conduct both GMP AAV upstream production by adherent and suspension culture.
Quality
Designed with QbD concept and optimization
Rigor
Yield
Reaching the maximum AAV yield up to 1E+15GC-1E+17 GC/batch
Cleanliness
Speed
Single-use technology and water supply to ensure batch cleaning verification
PackGene can provide cGMP AAV manufacturing for your project. Please provide your project details to get a quote or technical support from PackGene.
Resources
What is the source of PackGene cell bank?
PackGene’s h293 cell bank is officially authorized for commercial use.
What is the AAV output for a single batch fermentation?
PackGene offers single batch fermentation at several volumes, including: 2L, 7L, 25L, 50L, and 200L. AAV yields for each of these production volumes varies across AAV serotypes. As an example, AAV9 is a medium to high-yielding serotype, and expected yields for AAV9 are as follows:
| Expected yield for AAV9 | |
| Volume | Yield |
| 2L | 1E+14GC |
| 50L | 1E+16GC |
| 100L | 2E+16GC |
What method is used to determine the empty shell rate for AAV samples?
Viral empty shell rate can be determined using several techniques including anion chromatography HPLC, Analytical Ultracentrifugation (AUC), Transmission Electron Microscopy (TEM), CyroTEM, or VG Titer/Capsid titer. AUC, TEM and CyroTEM are typically not suitable for quantitative quality control determinations and thus PackGene’s standard method for empty shell rate determination is anion chromatography HPLC. PackGene can provide additional CyroTEM and AUC analytical services to serve as a secondary verification of the results derived from anion chromatography HPLC.
How are GC/ml and vg/ml related to one another, and how does PackGene determine GC/ml for AAV products?
The terms genome copies per ml (GC/ml) and viral genomes per ml (vg/ml) are interchangeable and equal in most cases. At PackGene we may test GC by both qPCR and ddPCR. Testing by qPCR involves the use of a calibration standard while ddPCR may use optional reference products. Measurements by qPCR are more likely to be influenced by inter-lab and inter-operator variables, and ddPCR generally shows lower %RSD precision. Typically, GC is determined by qPCR during process exploration phase and for intermediate products while the GC of final products is more often determined by ddPCR.
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