Currently, the R&D of ADC is on the eve of outbreak. However, many ADCs behind the marketed drugs terminated clinical trials due to safety or effectiveness issues. Therefore, the development trend of ADC drugs, clinical considerations, and verification guidelines may put higher requirements for the quality attributes and process development of ADC in the future. Major pharmaceutical companies must pay attention to the impact of unconjugated small molecule drugs on dose/exposure, unconjugated antibodies (affecting the relationship between total antibody levels and total ADC levels), and potential immune responses of ADC molecules. BOC Sciences can provide various ADC downstream purification technical support to meet the purification requirements after ADC coupling in large-scale production, which has a good effect on controlling the DAR value and removing impurities such as aggregates, unconjugated antibodies and small molecules.
Aggregates, unconjugated small molecule drugs and related impurities, DAR distribution and unconjugated antibodies are the impurities that need to be focused on in ADC drugs. The aggregation problem is one of the main obstacles in the development of ADC drugs, which may occur in every process of production as well as during transportation and long-term storage. In addition to triggering an immune response, aggregates can also alter the PK/PD of the drug. The causes of aggregation include high temperature, shear force, organic solvents, and the hydrophobicity of small molecule drugs. In addition, the bystander effect of ADCs relies on nonpolar and uncharged small-molecule drugs, and the excessive hydrophobicity of small-molecule drugs and changes in the conformational stability of antibodies after conjugation make ADC drugs easier to aggregate. Unconjugated small molecules will increase systemic exposure toxicity and affect the therapeutic window and safety of ADC drugs. Molecules with different DAR values may have different efficacy, safety and PK. Improving the homogeneity of ADC drugs is the development trend of ADC, which requires fixed-point coupling technology and necessary control of DAR value after coupling. The unconjugated antibody will occupy the drug binding site and affect the efficacy of the ADC. Therefore, detection and purification methods with high sensitivity and specificity are necessary for the cGMP production of ADC drugs.
BOC Sciences has been committed to exploring the purification process of ADC drugs, striving to provide comprehensive technical and service support for the downstream development of ADC drugs. Our purification processes include tangential flow filtration (TFF), ultrafiltration diafiltration (UFDF), size exclusion chromatography (SEC), hydrophobic chromatography (HIC), hydroxyapatite chromatography (HA) and membrane chromatography (MC).
The UFDF technique of tangential flow filtration has been widely used for ADC drug purification. The yield of the ultrafiltration or diafiltration technique for ADC purification can also be maintained above 90%. Studies have shown that small-molecule impurities, such as organic and conjugate impurities, have a lower molecular weight than the membrane pore size. Therefore, pH, membrane loading, transmembrane pressure difference, and flow rate during the filtration process have little-to-no effect on removing small molecular impurities.
SEC is a chromatographic strategy for separation and purification based on different molecular weights, which plays a crucial role in the purification of ADC drugs. SEC can play a role in buffer exchange, removal of aggregates, removal of non-protein impurities, and removal of small molecular impurities. In addition, the related chromatographic conditions are relatively mild, including neutral pH buffer and room temperature, which are beneficial for ADC drug stability maintenance. Generally, the SEC purification requires the selection of media with minimum interactions for process development.
In addition to small molecular impurities and aggregates, separation of different DAR species is another important goal of ADC purification. SEC is not suitable as a chromatographic technique for DAR separation due to its low resolution. As a non-denaturing condition analysis, HIC is often used for DAR determination of ADCs. In addition to its use in DAR assays, HIC can also be used to purify ADCs. Results from multiple studies demonstrated that HIC-purified site-specific ADCs retained both comparable cytotoxic activity and the desired target selectivity.
Besides HIC, another method for ADC purification under native conditions is IEC. IEC is mostly used in protein analysis, such as charge variation analysis, glycoform analysis and protein structure change analysis. Typically, IEC can achieve high recoveries while removing non-antibody-related impurities including small molecules and endotoxins, making it a promising method for ADC purification.
HA is a useful method for monoclonal antibody purification. The use of HA can reduce antibody aggregates from 60% to 0.1%. Therefore, HA was used after protein A chromatography. The protein A eluate contains purified antibodies and residual impurities associated with cell culture and leached protein A. Presently, HA chromatography can effectively remove impurities in ADC.
MC has higher capture efficiency and productivity than traditional resin chromatography, presenting the most promising applications in the recovery, separation, and purification of biomolecules. Due to the advantages of convective solute mass transfer within the adsorption membrane, MC has the potential to operate at much higher flow rates than packed columns, which can reduce the degradation and denaturation of biomolecules, as well as the buffer volume required for a given purification step. In downstream processes, MC is commonly used to remove aggregates and residual drug linkers from ADC components and to perform DAR species separation.
In addition to ADC purification support, BOC Sciences has cGMP manufacturing facilities in the US and UK, providing antibody modification, cytotoxin and linker chemical synthesis, and ADC conjugation and filling services for your ADC development. We have 40,000 square feet of cGMP production space and are equipped with multiple pharmaceutical (DP) filling lines. Our professional ADC development team has been operating strictly to ensure product quality. A professional project management team is also extensively activated to ensure on-time delivery of products and long-term after-sales handling. If you are interested in our ADC purification, please contact us for more information.
BOC Sciences' cGMP manufacturing can prepare ADCs for scale-up and commercialization. We have strict cGMP manufacturing standards, including scientific isolation controls, rigorous cleaning and sterilization processes, and FDA-compliant production and purification facilities.