Transforming Biomanufacturing: Potential of Cell Free Systems

Overview of Cell Free Systems

Cell-free systems are ­in-vitro platforms which allow occurrence of biochemical reactions in the absence of living cells. These expression systems utilize bio machinery harvested from the lysate of disrupted cells for the manufacturing of a wide array of macromolecular and small molecule products. Biomanufacturing utilizing cell free systems is an emerging area of research that enables the synthesis of different biomolecules, such as cytotoxic proteins, fusion proteins, post translationally modified proteins, antibodies, enzymes, vaccines and other complex proteins, without the use of living cells.

Types of Cell Free Systems

These systems can be broadly categorized into two classes:

• Reconstituted systems: These types of cell free systems are prepared using various factors which are essential for protein synthesis, including tRNAs, purified enzymes, amino acids, ribosomes, and energy molecules.  Such systems provide control over the components used for the synthesis of target products making it easier to manipulate the contents according to specific needs. Additionally, these types of cell free systems do not contain proteases and nucleases as crude cell lysates, which further helps in improving the production of many proteins. It is worth mentioning that the first reconstituted system, Protein Synthesis Using Recombinant Elements (PURE), was developed in 2001.
• Crude cell lysate-based systems: These types of systems consist of biocatalysts derived from cell lysis, and are commonly prepared from Escherichia coli, rabbit reticulocytes, Saccharomyces cerevisiae, wheat germ, and insect cells. It is worth mentioning that crude cell extract based cell free systems are less expensive and provide higher yields as compared to reconstituted systems.

Cell Free Biomanufacturing

The process of cell-free biomanufacturing involves a series of steps briefly described below:

1. Preparation of Cell Extract by extraction of cellular machinery from living cells
2. Preparation of Reaction Mixture by addition of co-factors and other essential components to the cell extract, creating an optimized environment for biomanufacturing
3. Addition of Template such as DNA or RNA for synthesis of desired biomolecules
4. Purification of synthesized biomolecules to facilitate further applications

Benefits and Challenges Associated with Cell Free Biomanufacturing

Cell free biomanufacturing is an emerging platform with various potential applications; however, a few associated constraints may pose a hurdle for its adoption. Key advantages and challenges associated with cell free biomanufacturing are given below.

Applications of Cell Free Systems

In the healthcare industry, cell free systems are primarily being used for the production of therapeutically important biologics, including membrane proteins, toxic proteins, monoclonal antibodies (MAbs), antimicrobial peptides (AMPs), vaccines, as well as high-value small molecule therapeutics.

In addition to the healthcare industry, other industries, such as food industry, cosmetics industry and enzyme synthesis industry, are also benefiting from the cell free manufacturing process.

Future Perspective

Since the introduction of such systems in the early 1900s, the platform has evolved considerably, with several technological advancements allowing the use of such systems for industrial production. Further, other improvements, including efficient production cycles, use of low-cost reagents and development of purified component platforms, have made cell free biomanufacturing a preferred option for synthesis of various types of biologics and small molecules. Although such systems are still plagued with various challenges, these systems continue to improve at a rapid pace. It is worth highlighting that companies, such as Nuclera Nucleics, Liberum Bio and Tierra Biosciences, have developed their own on-demand systems, suggesting that the use of cell free systems will continue to address the bottlenecks stemming from conventional biomanufacturing platforms.

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