The Rocky Road From Fed-Batch to Continuous Processing With E. coli

Abstract

Escherichia coli still serves as a beloved workhorse for the production of many biopharmaceuticals as it fulfills essential criteria, such as having fast doubling times, exhibiting a low risk of contamination, and being easy to upscale. Most industrial processes in E. coli are carried out in fed-batch mode. However, recent trends show that the biotech industry is moving toward time-independent processing, trying to improve the space-time yield, and especially targeting constant quality attributes. In the 1950s, the term "chemostat" was introduced for the first time by Novick and Szilard, who followed up on the previous work performed by Monod. Chemostat processing resulted in a major hype 10 years after its official introduction. However, enthusiasm decreased as experiments suffered from genetic instabilities and physiology issues. Major improvements in strain engineering and the usage of tunable promotor systems facilitated chemostat processes. In addition, critical process parameters have been identified, and the effects they have on diverse quality attributes are understood in much more depth, thereby easing process control. By pooling the knowledge gained throughout the recent years, new applications, such as parallelization, cascade processing, and population controls, are applied nowadays. However, to control the highly heterogeneous cultivation broth to achieve stable productivity throughout long-term cultivations is still tricky. Within this review, we discuss the current state of E. coli fed-batch process understanding and its tech transfer potential within continuous processing. Furthermore, the achievements in the continuous upstream applications of E. coli and the continuous downstream processing of intracellular proteins will be discussed.

Keywords: E. coli; burden reduction; continuous processing; from batch to continuous manufacturing; process understanding.