FAQ : Plasmid Manufacturing Service

FAQ Plasmid Manufacturing Service

Introduce your request using our contact us form. A first ballpark estimation will be rapidly sent to you. Based on the requested plasmid quantity and quality grade, a detailed quotation will be edited including the adapted strategy, the successive work packages and deliverables, the timing as well as the associated budget.

The turnaround time for the manufacturing of one R&D grade batch ranges between 4 and 6 weeks depending on the existence or not of a cell bank. The timing required for the manufacturing of a set of 4 plasmids will take between 9 and 11 weeks. Ten additional weeks are necessary for the release of one GMP batch.

The maximum quantity of plasmid that can be delivered is 20 g regardless of the quality grade considered (R&D, HQ or GMP). (Voir tableau dans section “”). We also offer a pDNA amplification service for quantities up to 50 mg at a discovery grade. (link towards FAQ Plasmid engineering)

A project of plasmid production can be performed using either a cell bank generated by Xpress Biologics or a cell bank provided by the customer. For biosafety purpose, the bank provided by the customer must be supplied with a CoA attesting the absence of bacteriophages.

HQ grade plasmid can be used as a drug substance for preclinical trials or as starting material for preclinical and clinical phases I/II trials. (Voir figure dans section “overview”). The produced and purified plasmid is characterized using a complete set of quality controls covering attributes related to physical state, identity, content, purity/impurity and safety. (Voir tableau ci-dessous: List of quality controls for the different quality grades of plasmids).

The GMP grade is required when the plasmid is used as a drug substance for all clinical trials and as starting material for clinical phases II/III trials. (Voir figure dans section “overview”). The produced and purified plasmid is characterized using a complete set of quality controls covering attributes related to physical state, identity, content, purity/impurity and safety. (Voir tableau ci-dessous: List of quality controls for the different quality grades of plasmids). The majority of the tests are compendial and therefore . Qualification of some specific tests such as the plasmid topology by HPLC has to be considered and is proposed by Xpress Biologics.

Inverted Terminal Repeats or ITR sequences that are present in AAV vectors are needed to guide genome replication and packaging during vector production. The ITR sequences usually form hairpin structures, which modifies the charge density of the plasmid leading to a different chromatographic behavior potentially decreasing the efficiency of the purification process. This is due to the fact that the presence of the ITR sequences renders the charge density of the supercoiled (SC) form similar to that of the open-circular (OC) form. The chromatographic conditions have been therefore modified in order to successfully separate both isoforms and allow the recovery of a proportion of SC form conform to the specification (˃95%).

The presence of repetitive sequences (ex: LTR sequences in LV vectors) could promote recombination events during the amplification phase. Xpress Biologics has developed a fermentation process avoiding such recombination events while allowing a plasmid yield at harvest compatible with industrial and economical constraints. Moreover, our analytical methods allow the complete sequencing of plasmids containing repetitive sequences and therefore can verify the absence of potential recombination.

Over the last 3 years, Xpress Biologics has managed around thirty manufacturing projects of plasmids dedicated to LV production, which corresponds to more than fifty batches, including R&D, HQ and GMP grade lots.

Over the last 3 years, Xpress Biologics has managed around twenty manufacturing projects of plasmids dedicated to AAV production, which corresponds to thirty batches, including R&D and HQ grade lots.

Xpress Biologics has adapted its manufacturing process (mainly fermentation) in order to take into account the presence of the long poly(A) tail in the plasmid sequence, tail up to 120 nucleotides being successfully produced and purified. When linearization is requested by the customer, a feasibility study is proposed in order to optimize the digestion conditions, with the aim to limit the amount of restrictive enzyme and therefore the production budget.

Due to the high degree of purity achieved following the purification protocol developed by Xpress Biologics (absence of nucleases), the stability of the plasmids appears to be excellent not only when the plasmid is frozen at -20 or -80°C (stability of more than 3 years), but also when stored at 4°C or RT. Moreover, based on the extensive experience acquired in recent years, Xpress Biologics can advise the customer on the nature of the formulation buffer to be used to ensure long-term stability. If required to support for instance the submission of a dossier for the introduction of a new drug, an extended stability study can be initiated following the ICH guidelines.

Polyplus offers a service of plasmid engineering based on an innovative assembly technology (e-Zyvec®). (Lien vers la page “Plasmid Engineering Service”). Online software and dedicated plasmid experts can help you to define the plasmid features that fit to your application and that are compatible with large scale manufacturing requirements.