From research to commercialization: what are the changes of scale and quality ?
Each stage involving the use of mAb , from research to industrial production, involves significantly different quantities and quality of antibodies. The requirements for purity and characterization are obviously increasing and maximum during use in humans.
While a few milligrams are sufficient for studies at the research stage, it is not uncommon to produce several hundred kg of the same antibody intended for certain oncological indications.
In terms of the processes for preparing recombinant proteins, there is generally a distinction between upstream and downstream platforms , respectively dedicated to cell culture and purification activities.
The upstream platform
It includes all the stages which go from the culture of the clone producing the mAb to the separation of the cells and the culture medium produced by the last bioreactor.
By extension, the preliminary steps including the generation of the cell line (after transfection with the vector of interest), the selection of the clone and the creation of the cell bank can be included in the upstream platform .
The selection of a clone, the optimization of culture conditions and the change of scale (from a few milliliters to several thousand liters) require real know – how. With the expertise and optimization cycles during the development phases, it is not uncommon to obtain cellular productivities 5 to 10 times higher than that of the initial clone.
The economic advantage is then undeniable for the implementation of the process at the industrial stage. Whatever the degree of optimization, the challenge, before moving on to industrialization, will be to demonstrate mastery of the process while ensuring the final quality of the product.
Production technologies have evolved significantly in recent years, even if the principle of cultivation in a bioreactor remains the same. The emergence of single-use systems has greatly facilitated the work of developers and producers. It is now conceivable to carry out in a single-use system all the stages of cell culture from the thawing of the cell tube of the cell bank to the bioreactor of 1000 or 2000 liters.
These bioreactors are equipped with a sterile plastic bag, a stirring system and all the “connectors” for controlling cell culture and transferring media. At the end of culture, the cells are recovered, the bag is replaced by a new one, allowing the bioreactor to be quickly operational again. Most of the peripheral equipment of the bioreactor (media preparation, connectors, filtration, etc.) is also designed in a single-use form.
The absence of sterilization, cleaning and cross-contamination are the main advantages of these systems. The current capacity limit for bioreactors (2,000 L) is compatible with the production of batches of antibodies intended for clinical trials or for small therapeutic markets (niche markets, requiring production not exceeding a few kg).
In the context of the commercial production of antibodies intended for the oncological field for example, the use of bioreactors of more traditional design and of large capacity remains required (up to 20,000 liters). The reader will be able to refer to a recent review dealing in depth with the specifics of the upstream platform .
The downstream platform
It includes all of the stages making it possible to obtain a purified antibody from the clarified culture medium (rid of cells). The active ingredient obtained must then be treated according to an appropriate formulation before being bottled for clinical trials or marketing.
In the case of antibodies, the standard purification methods most often include an affinity chromatography step (on protein A) followed by two chromatography steps by ion exchange or by hydrophobic interaction. All the methods implemented contribute to the elimination of cellular contaminants or those brought by the process.
In particular, contaminating proteins of the host cell, DNA, protein A or viruses potentially brought by the cell line must be eliminated or limited to concentrations below regulatory standards. In the case of viruses, elimination or inactivation by at least two orthogonal steps in the process is a regulatory requirement.
The purity objective is accompanied by an objective of optimal performance of the platform. Depending on the choices made by the developer and according to the intrinsic characteristics of the antibody, the purification yields generally encountered are from 60 to 70%. With the dramatic improvement in cellular productivities, purification has become the bottleneck in the production of antibodies.
The downstream stages represent 60 to 70% of the total cost of production of the antibodies. Efforts to simplify this platform and reduce costs have become major challenges.
The development of alternatives to protein A , the creation of new resins or new loaded filtration membranes will provide competitive solutions. Similarly, the emergence of single-use technologies in the downstream platform (filtration, preparation of buffers, connectors, etc.) makes it possible to reduce process times while ensuring the reliability of the quality of the active ingredient.
The rapid availability of antibodies for clinical trials is absolutely essential in the development cycle and the prospect of commercial success. In parallel with the upstream and downstream platforms , analytical methods are implemented to characterize the antibody of interest. Given the increasing demand for purity and characterization, analytical techniques are now available that push the limits of quantification of the subtle variations observed in the structure of antibodies ever further.
As many of the articles in this issue attest, antibodies are major molecules therapeutically. Many pharmaceutical entities have entered this promising niche. Competition is therefore active, all the more so since the therapeutic targets targeted are much fewer than the hundreds of antibodies in development. On the other hand, the high costs of research, development and production of antibodies have made them expensive therapies.
Reduced reimbursement policies and the emergence of biosimilars pose a new challenge to the pharmaceutical industry. The financial risks and the excitement of competition stimulate minds in search of faster, more reliable and more economical technologies.
Cellular productivity has literally exploded in the past decade. This parameter partly responds to the economic aspects because it simply makes it possible to reduce the volumes of bioreactors required to obtain an equal quantity of antibodies. If the upstream advantage is obvious, everyone agrees today on the need to find new solutions to the downstream platform .
This is one of the immediate challenges that the biopharmaceutical industry will face. But right now, with the number of antibodies in development and the technological systems in place, this same industry offers optimistic prospects for bringing patients new therapeutic biomolecules.