Beschreibung:
<jats:title>Abstract</jats:title>
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<jats:title>Background</jats:title>
<jats:p>Significant challenges exist in downstream purification of bispecific antibodies (BsAbs) due to the complexity of BsAb architecture. A unique panel of mispaired species can result in a higher level of product-related impurities. In addition to process-related impurities such as host cell proteins (HCPs) and residual DNA (resDNA), these product-related impurities must be separated from the targeted BsAb product to achieve high purity. Therefore, development of an efficient and robust chromatography purification process is essential to ensure the safety, quality, purity and efficacy of BsAb products that consequently meet regulatory requirements for clinical trials and commercialization.</jats:p>
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<jats:title>Methods</jats:title>
<jats:p>We have developed a robust downstream BsAb process consisting of a mixed-mode ceramic hydroxyapatite (CHT) chromatography step, which offers unique separation capabilities tailored to BsAbs, and assessed impurity clearance.</jats:p>
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<jats:title>Results</jats:title>
<jats:p>We demonstrate that the CHT chromatography column provides additional clearance of low molecular weight (LMW) and high molecular weight (HMW) species that cannot be separated by other chromatography columns such as ion exchange for a particular BsAb, resulting in ≥98% CE-SDS (non-reduced) purity. Moreover, through Polysorbate-80 (PS-80) spiking and LC–MS HCP assessments, we reveal complete clearance of potential PS-80-degrading HCP populations in the CHT eluate product pool.</jats:p>
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<jats:title>Conclusions</jats:title>
<jats:p>In summary, these results demonstrate that CHT mixed-mode chromatography plays an important role in separation of product- and process-related impurities in the BsAb downstream process.</jats:p>
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