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Process Development & Characterization

A commercial-scale biologic production is only possible when bench-scale prototypes are scalable to pilot-scale and high-volume bioreactors. Parameters should be established both at the shaker flask scale and at the larger scale reactors so that wastage of valuable resources and time are saved. Therefore, it is critical to establish the production parameters following rigorous experimental design and verification of as many variables as feasible. Process development must also include parameter testing at various bioreactor volumes.

  • Upstream Process Optimization
  • High productivity with > 6 g/L titer
  • Cell viability > 90%
  • Supporting bioanalytic to ensure quality

Upstream Process Flow

 

  • Use of nonproprietary commercially available media and feed for screening studies allows transfer to any CDMO (Contract Development and Manufacturing Organization)
  • Media and feed screening in Shake Flasks
  • Process parameter screening and optimization in 1L Bioreactor
  • Process consistency and scalability happens in 5L Bioreactor

Ambr250® for Process development

  • Ambr250® to support Process development & Late-stage Clone evaluation: Ambr250® is a bioreactor system for parallel fermentation or cell culture development with intuitive programming and maximum flexibility
  • Fully automated Ambr250® 24 units, integrated to NOVA Flex-2 & Vi-Cell
  • Culture Performance
    • Metabolites, pH, DO and CO2
    • VCD and %Viability
  • Operating Parameter
    • Multifactorial DOE to screen process parameters like- pH, DO, Temp and agitation
  • Process parameter 
    • Optimization and identifying the best operative parameters

Process Development (PD) Case Studies: PD to Increase titer & Improve Product Quality

  • Improved titer from 20 to 300ug/mL with >85% fully formed protein.

  • Media adaption
  • Shake flask fed batch evaluation with different media and Feed.
  • Scalable and consistent data from shake flask to 1L and 5L Bioreactor in terms of VCD and titer profile
  • Glycan profile, SEC (Size Exclusion Chromatography), cIEF, Mass Spec, SPR (Surface Plasmon Resonance) is comparable to Originator Anti-inflammatory Tumor Necrosis Factor Inhibiting Agent 

Part A: Media Adaptation; Growth Profile

  • Master Cell Bank (MCB) received from a client was thawed, scaled up in the original medium, Media A, based on Aragen Bioscience protocol.
  • SP2/0 cells were adapted in different mediums. The SP2/0 adapted well (viability >90%) with Media B, D and F. These media were used in the next stage.

 

Downstream Process Development (DSP)

The design of manufacturing process that consistently ensures the safety and efficacy of the drug is critical for biopharmaceuticals development. Hence, a downstream purification process is crucial throughout the lifecycle of a product to ensure product quality. Early process development phases are driven by the requirement to expedite clinical development of a biotherapeutic towards clinics. The DSP can be challenging as a variety of downstream separation techniques and combinations are considered. Furthermore, each of the unit operations depends on many process parameters and material attributes that need to be optimized and validated for robustness. Often an experimental investigation of all parameter combinations is challenging.

Typical key steps of DSP

  • Harvest and Filtration-Depth filtration, Hollow Fibers
  • Primary Capture- Affinity and Ion exchange
  • Viral inactivation- Low pH
  • Buffer Exchange and concentration
  • Formulation

Risk Ranking for Polishing Steps

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