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Structural Characterization

Structure determination

VelaLabs has expanded its physico-chemical portfolio via the implementation of a range of diverse structure determination methods.

Sophisticated Structure Determination Methods at VelaLabs

VelaLabs offers also liquid chromatography mass spectrometry (LC-MS), gas chromatography mass spectrometry (GC-MS), size exclusion chromatography multi-angle laser light scattering (SEC-MALLS) and capillary electrophoresis (CE). Subsequent to the acquisition of our new equipment and integration of this innovative technology, we are now offering modern, up-to-date analytical methods for determining structural characteristics of peptides, proteins and small molecules.

Sciex Triple TOF 6600

Mass Spectrometry of Proteins: An Innovative Era in Characterizing Biological Therapeutics

Mass spectrometry (MS) is a rapidly-emerging technique to characterize protein drug therapeutics. Such proven analytical methods are now routinely used to characterize biosimilars, biopharmaceuticals & biological products during pre-clinical and clinical development as well as analysis of products during process development. The new portfolio at VelaLabs that encompasses protein mass spectrometry consists of three consecutive stages of implementation – each dedicated to support our clients in all phases of product development and before batch release.

Stage 1: Basic Mass Spectrometry Analysis of Your Protein

Here we are focused on establishing and offering fundamental mass spectrometry-based analyses that include:

  • Protein identification and confirmation via gel-based and gel-free methods plus screening of different clones for biosimilar products
  • Recombinant protein intact mass measurements to, e.g., differentiate individual protein glycoforms
  • Absolute, ‘pure’ protein quantitation via spiked ‘heavy’-labelled peptides of known concentration and comparison to unknown levels in specific biological samples

Agilent 1290 Infinity II

Stage 2: Characterize and Quantitate Your Protein

For projects that require a deeper level of analysis and/or for our clients that wish to continue further from Stage 1, we can offer additional analyses such as:

  • Complete protein sequencing whereby recombinant proteins are treated with multiple proteolytic enzymes and the generated peptides are analyzed by LC-MS
  • Identification and/or localization of post-translational modifications that are important in establishing the integrity of biopharmaceuticals, e.g., assessing the degree of oxidation, de-amidation, glycosylation, phosphorylation etc.and/or protein aggregate formation
  • Global relative protein quantitation of complex mixtures via labelled and label-free approaches
  • Structural relationships to understand and identify protein-protein, protein-small molecule interactions

Stage 3: Tailored Approaches Dedicated to Your Requirements

For even more advanced projects, we are available to discuss and design specific mass-spectrometry based proteomic approaches that will involve:

  • Case-by-case discussions based on client and project requirements
  • Scientific support during process development and validation
  • Comparability studies for biosimilars to ensure that batch-to-batch variability is minimized

These three stages will assist in several important phases of the industrial pipelines of our Clients:

Discovery Phase: Non-GxP proteomic approaches will aid in: (i) clone selection; (ii) monitoring/determining in vitro proteome-wide effects; (iii) absolute quantitation of biomarkers; (iv) characterization of mutant versus wild-type protein forms via intact mass analysis and de novosequencing; and (v) cross-validation comparison with a traditional portfolio (bioassays etc.)

(Pre-) clinical Studies: Here, LC-MS will provide an analytical portfolio that enables protein-based drugs measurements in complex matrices (e.g., human serum/plasma samples).  As a proof-of-principle, a simulated pharmacokinetic (PK) study on pegylated and non-pegylated granulocyte colony stimulating factor (pegG-CSF and G-CSF) and on a second biopharmaceutical, rituximab, is currently evaluated.

Market Authorisation: Protein mass spectrometry will be used to perform: (i) stability studies, e.g., determining the degree of protein oxidation and de-amidation of a biopharmaceutical product; and (ii) comparability studies between batches via peptide mapping and glycoform analysis.

Agilent 7100 CE

Batch Release: Via peptide mapping, LC-MS will be used to: (i) assess protein product batch-to-batch consistency; (ii) monitor/confirm recombinant amino acid sequences; and (iii) determine the degree of impurities, e.g., via host cell protein analysis. Similarly, intact protein analysis will confirm: (i) complete protein expression; and (ii) the major glycoforms and glycosylation pattern of the Protein.

Extractables and Leachables

VelaLabs also offers characterization of Extractables and Leachables (E&L) components from the primary packaging materials of (bio-) pharmaceuticals and biosimilars.

Via combination of gas chromatography mass spectrometry (GC-MS), high resolution liquid chromatography quadrupole time-of-flight (Q-TOF), and liquid chromatography mass spectrometry (LC-MS)

Extractables are investigated during material characterisation and during selection of different primary packaging materials. Leachable studies are required for market authorisation as compounds that have leached from the primary packaging material into the drug product must be investigated.

Via the combination of gas chromatography mass spectrometry (GC-MS) and high resolution liquid chromatography quadrupole time-of-flight (Q-TOF), and liquid chromatography mass spectrometry (LC-MS) we are implementing an orthogonal approach to ensure maximal coverage of volatile and non-volatile compounds. Unknown substances can be quantitated using an internal standard response method. For components where the levels exceed safety-relevant thresholds, additional structural information can be investigated to predict a possible structure. For specific analytes, absolute quantitation methods can be established and validated.

Objectives Extractables Accelerated Leachables Leachables
  selection of extractables to be monitored selection of leachables to be monitored relative quantitation using surrogate standards
  worst case scenario close to realityincluding reaction products with API identification of leachables above the analytical evaluation threshold (toxicological evaluation required)
Strategy solvent extraction of primary packaging material storage of drug product at higher temperatures storage of drug product for complete shelf life

Market Authorization

Extractable and leachable studies of primary packaging material for final drug products will enable our clients to select the optimal packaging material(s) and demonstrate safety for complete shelf life.

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