CreDA™ developability assessment service from Creative Biolabs is able to characterize the electrostatic properties of the candidates in terms of isoelectric point and charge profile, which can guide the selection of appropriate candidates for downstream development.
For protein purification, taking antibody as an example, protein A affinity chromatography is usually followed by polishing steps consisting of orthogonal methods such as anion exchange chromatography (AEC) or cation exchange chromatography (CEC) in flow-through mode. The use of these chromatographic methods imposes thresholds on the protein isoelectric point (pI). This arises from the fact that the electrostatic properties of the protein (together with the pH and ionic strength of the sample) will determine the protein affinity for ion exchange resins. For example, a pI high enough (e.g. pI>7) is considered a prerequisite for binding and nonbinding to cation and anion exchange chromatography media, respectively. Thus, it is of interest to select candidates that fulfill requirements of purification in an established downstream processing platform. Apart from the pI (the net charge), charge distribution of the biopharmaceuticals may also influence the solution behavior and product stability during development stages. Strong asymmetric charge distribution was shown to be correlated to a propensity of such molecules to self-interact, potentially leading to high viscosity or high aggregation tendency. Based on these considerations, our developability assessment service will characterize the electrostatic properties of the candidates in terms of isoelectric point and charge profile, which can guide the selection of appropriate candidate for downstream development.
Fig.1 Electrostatic properties assessment of the drug candidates.
The isoelectric point (pI) is the pH at which the protein has zero net charge, and it is highly dependent on the type of amino acid residues in the protein, the residue solvent accessibility and spatial arrangement. The pI value can be determined either in silico from the protein sequence or, more accurately, in vitro by isoelectric focusing (IEF) or capillary isoelectric focusing (CIEF).
The analysis of charge variants is a regulatory requirement for biotherapeutic proteins. These charge heterogeneous molecules can be the results of a variety of enzymatic post-translational modifications during manufacture, such as glycosylation and lysine truncation. In addition, chemical modifications occurring during purification and storage such as oxidation or deamination also contribute to the charge heterogeneity. Charge variants are identified by capillary zone electrophoresis (CZE) or cation-exchange chromatography. More detailed characterization of the variants is based on peptide mapping and LC-MS.
Electrostatic properties are key attributes that may influence the purification process of biotherapeutic proteins. Our CreDA™ platform supplies with the state-of-the-art analysis method, such as CIEF and iCIEF, to assess the electrostatic properties of your drug candidates, aiming to screen out the lead molecule most appropriate for downstream development. If you are interested in this service, please contact us for more information and a detailed quote.
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