To confirm the biosimilarity of a potential competitor biologic product to the original reference biologic, analytical
studies must be carried out to show that it is indeed highly similar to the reference product. In general, extensive comparative
physicochemical and functional studies are required.
Little is generally known about the originator company's manufacturing process as this is proprietary information. Variability
in the exact nature of the biologic product is inevitable, thus, its intrinsic similarity to the reference product must be
In addition, any impurities related either to the product itself or the process used to manufacture it must be identified,
characterized, quantified, and compared to those of the reference product as must all the substances deliberately included
within the product. The type, nature, and extent of any differences between the biosimilar and the reference must be clearly
described and discussed in the application for approval.
A number of different factors should be considered when assessing if the two products are indeed highly similar. First, the
expression system must be considered, including the identity of the cell line being transfected, the sequence of the transgene,
any promotors or other control regions, and the overall genetic identity and stability of the cloned gene. Biosafety testing
of the cellular substrates including the master cell bank (MCB) and cells at the limit of in-vitro cell age used for production must also be performed. The gene-copy number, and the gene-sequencing process, whether messenger
ribonucleic acid (mRNA) or complementary deoxyribonucleic acid (cDNA) should also be determined in the cellular substrates.
Fluorescence in-situ hybridization (FISH) analysis and restriction enzyme analysis should also be carried out. This comparison of the MCB and
the cells at the limit of in-vitro cell age (considered to be the 'worst case scenario' in a bioproduction setting) enables the demonstration of the genetic
stability characteristics of the MCB.
Next, the manufacturing process must be studied, including the cell line used, the glycosylation structure and heterogeneity
it produces as well as both product expression levels and the expression of endogenous retroviral particles. Also, the viability
and productivity of cells, product integrity, and degradation products and levels of host-cell protein and DNA must be assessed.
The nature of the media is also important, such as whether or not it contains serum, is protein-free or chemically defined.
Processing considerations, such as the bioreactor format and the downstream process, must also be considered, along with the
removal of process- and product-related contaminants, virus inactivation and removal, product formulation, and product stability.
Finally, as with all medicinal products, the viral and microbial safety of the biosimilar must be considered. These are assured
by three complementary approaches:
- Testing of starting materials for viral and microbial contaminants
- Testing process intermediates at appropriate stages in the manufacturing process for any contaminating viruses, mycoplasma,
bacteria, and fungi
- Analytical characterization.