What is a Biopharmaceutical?
A biopharmaceutical, also termed a “biologic”, is a medical therapeutic (or drug) that has been synthesized or extracted from a biological source, such as a cell.
Biopharmaceuticals vs pharmaceuticals
The main difference between pharmaceuticals and biopharmaceuticals is that pharmaceuticals have been chemically synthesized whereas biopharmaceuticals have been made using some form of biological source or host.
From March 2020, chemically synthesized polypeptides, such as insulin, will be considered as biologics by the FDA and drug submissions will be considered according to biologics regulations.
Some examples of biopharmaceuticals include:
- Proteins and glycoproteins, including antibodies
- Cells, such as stem cells
- Organs and tissues (e.g. transplants)
- Gene therapies
Structural and physicochemical characterization are a crucial part of biopharmaceutical product development. These are some of the analytical methods and tests that allow drug developers to be sure that they have correctly manufactured their target drug. The FDA and EMA have set out their own guideline expectations for the protein characterization methods to be performed. Both agencies have adopted the characterization approach outlined in a regulatory document called ICH Topic Q6B.
ICH Topic Q6B provides examples of technical approaches to be considered for structural characterization and confirmation, and evaluation of physicochemical properties of the desired product, drug substance and/or drug product. The specific characterization assays employed will vary from product to product and alternative approaches may be appropriate in many cases. The guideline recognizes that new analytical technology and modifications to existing technology are continuously being developed and should be utilized when appropriate.
ICH Q6B requires data covering the following structural and physicochemical elements for the biopharmaceutical:
|Analytical requirement||Methodology used for data provision||Instrumentation requirements|
|Amino acid sequence||N-terminal sequencing (Edman chemistry)
On line LC-MS (with MSe and/or MS/MS)
MALDI-MS and MS/MS
|Amino acid composition||Amino acid analysis||Amino acid Analyzer (e.g. RP-HPLC with fluorescence detector)|
|Terminal amino acid sequence||N-terminal sequencing (Edman or mass spectrometry)
Mass spectrometry based sequencing for the C-terminus
Mass spectrometer capable of providing N- and C-terminal sequence information
|Peptide map||Peptide mapping||On line LC-MS (with MSe and/or MS/MS)|
|Sulphydryl group(s) and disulphide bridges||Non-reduced peptide mapping||On line LC-MS (with MSe and/or MS/MS)|
|Carbohydrate structure||Monosaccharide composition analysis
Oligosaccharide population analysis
Glycosylation site analysis
LC and on line LC-MS (with MSe and/or MS/MS)
MALDI-MS and MS/MS
|Molecular weight or size||Intact molecular weight analysis||On line LC-MS
|Isoform pattern||IsoElectric Focusing (IEF)||Imaged Capillary IEF (icIEF)|
|Extinction coefficient||Optical density measurement and protein concentration determination||UV spectrophotometer
Amino acid Analyzer (e.g. RP-HPLC with fluorescence detector)
|Electrophoretic analysis||Charge and size based electrophoretic analysis||icIEF
Capillary Gel Electrophoresis (CE-SDS)
|Liquid Chromatography||Charge, size and hydrophilicity/hydrophobicity based chromatographic analyses||Ion-exchange chromatography (IEX)
Size Exclusion Chromatography (SEC)
Reversed Phase Chromatography (RP-HPLC)
|Spectroscopic profiles||Secondary and tertiary structure analysis||NMR (1D and 2D)1
Circular Dichroism (CD)
Fourier Transform-Infra Red (FT-IR)
Fluorescence (Intrinsic and Extrinsic)
|Analysis of truncated forms||Intact molecular weight analysis
Size based chromatographic and electrophoretic analysis
|On line LC-MS (with MSe and/or MS/MS)
|Analysis of other modified forms, including Post Translational Modifications (PTMs)||Intact molecular weight
|On line LC-MS (with MSe and/or MS/MS)
|Aggregation||Size based chromatography, Analytical UltraCentrifugation (AUC)||Size Exclusion Chromatography with Multi Angle Laserlight Scattering (SEC-MALS)
Sedimentation Velocity-Analytical UltraCentrifugation (SV-AUC)
1 HDX-MS data can also be used if NMR data is not available.
When Should I Perform Structural Characterization Studies?
The above analytical techniques should be used to characterize multiple batches of the product throughout the development process. It is very important to begin characterization studies at the early stages and scales of product development to ensure that the product and manufacturing process are correct from the outset. These studies form an ongoing investigation into the product through scale up and product processing to ensure that no changes are introduced during product manufacturing and process development. Data-led decisions can then be taken to modify procedures if necessary. The final aim of this is to build a package of data that will satisfy the regulator that the manufacturing process is robust and produces a consistent product.
The structural characterization methods should also be used to assess the product following any significant changes in the manufacturing process. These could include:
- improvement of the manufacturing process
- increasing the manufacturing scale
- change of manufacturing site
- making changes to improve product stability
- an unwanted or unexpected change
- g. pH or temperature deviation
What Characterization Methods Should I Perform?
Whilst the full list of analytical techniques is ultimately needed for detailed protein characterization, an appropriate group of selected techniques can be applied at the outset of the product development cycle. These select methods should be carefully chosen such that they will test the critical structural parameters of your biopharmaceuticals. Common examples are:
- Molecular weight analysis, for product intactness
- Peptide mapping, for basic sequence analysis and assessment of post-translational modifications
- Glycosylation population analysis for glycoproteins
- Aggregation analysis, using SEC-MALS and/ or SV-AUC
- Basic higher order structure, to check the folding is correct
How Do I Start Characterizing My Biopharmaceutical?
BioPharmaSpec scientists would be happy to discuss when and how to perform the correct characterization methods for your biopharmaceutical. Please contact us here.