Plasmid DNA is manufactured on an industrial scale by means of tank fermentation, using either batch or fed-batch methods with engineered strains of E. coli as the host organism. In a large scale process, it is critical to achieve the maximum yield of plasmid DNA (pDNA). This is necessary not only for the obvious economic reasons, but for quality (purity) and minimal environmental impact as well.
Process development starts by designing and optimizing the plasmid construct for optimal expression and production in the respective host cells. Assuming this has already been done (see NTC’s Vector Development Services), the next step is optimizing fermentation. Traditionally, yields of 10-100mg/L have been achieved, although the literature reports some results in the 200-250mg/L range.
However, more recent improvements in plasmid DNA processing at NTC (patents pending) have now made it possible to achieve phenomenal yields of 2g/liter, and greater. Factors affecting yield include: plasmid backbone (replication origin), host strain, metabolic burden placed on the host cell, insert stability (repeats, palindromes, etc.), as well as the process parameters themselves. NTC’s proprietary improvements in yield have dramatically decreased production costs and have routinely resulted in highly-purified, covalently closed circular monomer pDNAs, which also resulted in dramatically reduced waste streams and a cleaner environment. Today, NTC is the only company in the World offering such a wide range of services combined with stellar yields.
Managing Risks
Process development at NTC is accomplished by determining the optimum methods for achieving at least 300mg-2g/L pDNA production in a scalable fermentation system and by transferring the optimized process to the production facility. This involves a standard license agreement, with the majority of technology acquisition costs postponed until success is achieved in clinical trials and pending FDA approval. Furthermore, the price of process development is based on the actual yields achieved with your plasmids, so the risk is mitigated by success. Once a successful process is devised, additional changes in the customer’s DNA backbone can generally be made and scaled-up with minimal effort.
Return on Investment
The rationale for performing Process Development at the early stage (prior to commencing clinical trials) is that: 1) the World’s leading Process can be locked down (i.e., you should not have to repeat preclinical or clinical trials due to last minute changes in the Process); 2) the Process can be transferred to a GMP facility (or in-house production suite) at reasonable cost with the necessary IP in place; and 3) the cost will not escalate due to forced acquisition of technology at a late time point (‘buy before you try’). The tremendous gain in productivity will pay for itself many times over compared to a locked-down, sub-optimal Process.
