Critical Quality Attribute (CQA) analysis
The development and manufacturing of biologics necessitate robust, reproducible product characterization throughout all stages of the drug process development. It is important to identify and understand your drug product’s Critical Quality Attributes (CQAs) – i.e., the physical, chemical, biological, and microbiological characteristics affecting your product’s quality. CQA information enables faster process and control strategy development and increases product and process knowledge. Furthermore, it allows you to progress into clinical trials and regulatory approval submissions confident of your drug’s safety, quality, and efficacy.
Mass spectrometry (LC-MS) enables fast, reproducible, quantitative evaluation of product-specific attributes and process-related impurities. Our services include intact mass analysis and peptide mapping to provide a detailed characterization of your drug substance (DS) for use in process and control strategy optimization. Furthermore, you’ll get a thorough overview of your DS’s stability, function, and purity.
Reproducible, quantitative intact mass and peptide mapping of CQAs enable
Detailed product and process understanding
Optimize the production process and evaluate the impact of formulation and manufacturing changes on product quality
Ensure the quality of your product throughout its lifecycle
Develop control strategies for monitoring product characteristics affecting product quality, and reduce the risk of variability in the final product
Faster regulatory approval
Increase transparency and understanding of your product’s chemistry, manufacturing, and controls (CMC) for expedited processing by regulatory authorities
The ICH Q6B guidelines call for thorough protein structure characterization of biologics and assessment of drug substance heterogeneity. You must demonstrate protein and batch consistency during drug development, following process or manufacturing changes, and after up-scaling or contract manufacturing organization (CMO) tech transfer. Early, pre-clinical CQA analysis helps prevent delays due to stability issues or other challenges during the clinical trial stages.
Intact mass analysis and peptide mapping services using LC-MS are highly sensitive and selective techniques for determining structural characterization and post-translational modifications (PTMs), which may affect the quality and efficacy of your final drug product. LC-MS data provides strong evidence of correct expression of specific proteins, protein impurity clearance, and product stability for regulatory submissions and product release testing.
Glycosylation is a typical PTM whose presence or absence, sites of attachment, and relative abundance can significantly impact the efficacy, stability, and activity of proteins and monoclonal antibodies (mAbs). We can compare batches, originator and biosimilar, or your process development samples using intact mass analysis and/or a complete glycan characterization - including N-glycan analysis.
Our LC-MS-based quantitative peptide mapping services also allow a thorough characterization of other PTMs, such as deamidation, oxidation, methylation, and pyro-Glu.
For many proteins and mAbs, correct disulfide linkage is critical for optimal biologic function. Product stability and activity issues often correlate with an abnormal pattern of disulfide bonds which affect the protein conformation and structure.
For this reason, the ICH Q6B guidelines call for analyzing disulfide bonds and free cysteines in biopharmaceuticals.
Using LC-MS, we perform protein mapping to determine the number and expected positions of disulfide bridges and free sulfhydryl groups in both non-reduced and reduced proteins and mAbs.
To avoid significant setbacks during regulatory approval, you must have a complete picture of your protein product’s CQAs and use the optimal analytical assays for each attribute to document compliance with regulatory requirements.
Combining different types of chromatography with our mass spectrometry services for intact mass analysis and quantitative peptide mapping provides you with the most comprehensive data available for your documentation.
Typical project process
You typically work with
We like to start with an online meeting to learn more about your project. Based on your needs and details about your samples, you will receive a draft proposal outlining the suggested analyses and expected timeframe.
After signing the final project proposal, we will contact you for information about shipping samples. We will inform you of an estimated report delivery date as soon as we receive your samples.
A project leader will oversee the project and send you status updates by email at regular intervals.
The analysis varies according to the project but typically includes:
- Intact mass analysis performed by LC-UV-ESI MS.
- Reduction/alkylation, protease digestion, and LC-ESI MS/MS peptide.
- Disulfide bond mapping by non-reduced peptide mapping analysis.
- Quantification of major post-translational modifications.
- N-glycan analysis.
You will receive the analysis report by email. Depending on the project, it includes:
- Objectives, description of analytical procedure, results, and conclusions.
- Selected raw data, e.g., excel sheets. Additional raw data may be provided to you upon request.
Upon project completion, your team is invited to review the results at an online meeting.
Overview of CQA analyses according to ICH Q8B
|Quality attribute (QA)|
Amino acid sequence coverage
LC-MS intact mass
Peptide mapping (1-3 different enzymatic digests)
From intact mAb
Map of N-glycosylation
Release N-glycan analysis
|Product heterogeneity, related substances, and impurities||
N- and C-terminal modifications
Charge and size heterogeneity
IEX MS or SEC/RPLC MS (incl. de-glycosylation)
|Chemical modifications||Position of glycation||
Peptide mapping (trypsin digest plus extra if needed)
IEX MS with de-glycosylation
Alternatively, RPLC MS or SEC MS with de-glycosylation
|Secondary structures||Disulfide linkages||Peptide mapping (non-reduced)|
What clients say
CMC Project Leader, R&D
UK biopharma company
Director, Protein and Analytical Chemistry
Research-based pharmaceutical company
Dorrit Andersen, Regulatory Affairs Consultant
DTA Consulting, Denmark
planning the study”
VP CMC and Quality
Kristiina Hyvärinen, Director QC, viral products
Targovax ASA, Finland
Torben Lund-Hansen, PhD, SVP
Head of Technical Operations
Y-mAbs Therapeutics Inc., USA
Head of CMC, C> Division
GTP Bioways, France
Why combine intact mass analysis and peptide mapping services for finding critical quality attributes?
Intact mass analysis is superior for identifying and quantifying different modifications in your intact drug molecule, whereas peptide mapping locates each modification in detail. Combining the two analyses allows e.g., with intact mass analysis detect that a molecule has 10 % oxidation, followed by peptide mapping to identify the exact site of the oxidations.
Why should I identify the CQAs in my drug molecule?
Knowing the CQAs can help you foresee potential problems and issues that may influence the stability and activity of your drug product. Monitoring the CQAs is essential for consistency of the manufacturing process and comparability of DS batches, as well as regulatory documentation
What are the typical CQAs of protein biologics?
Typical CQAs can be specific glycoforms, charge variants (e.g., deamidation), easily oxidized sites, and aggregations.
Talk to us
Whatever protein-related challenge or question you may have, we would love to help. Our experts can help you decide on the best analytical approach for your project by email or online meeting - providing advice without obligation.