Seattle Genetics, Inc.

Seagen Inc.

John Fay Valliere-Douglass

Oscar Salas

Damon L Meyer

Robert Lyon

Kevin Beam

Bradley Hayes

The present invention provides methods and systems for the rapid determination of the intact mass of non-covalently associated antibody heavy chains (HC) and light chains (LC) which result from the attachment of drug conjugates to interchain cysteine residues. By analyzing the antibody-drug conjugate (ADC) using native desalting conditions, the intact-bivalent structure of the ADC, which ordinarily would decompose as a consequence of denaturing chromatographic conditions typically used for LCMS, is maintained. The mass of the desalted ADC is subsequently determined using desolvation and ionization ESI-MS conditions. The methods described herein provide for direct measurement of the intact mass of an ADC conjugated at interchain cysteine residues. The methods described herein also provide for the relative quantitation of the individual ADC species.

Intact mass determination of protein conjugated agent compounds

The invention is based in part on the observation that a CHO cell oxidizing enzyme, particularly QSOX1, can survive a seemingly rigorous antibody purification process to reduce subsequent conjugation efficiency of the antibody to a drug. Whether the oxidizing enzyme survives the purification procedure depends on which purification techniques are employed which can vary from one antibody to another. With knowledge that contamination with a CHO cell oxidizing enzyme is a potential problem for subsequent conjugation, a suitable purification scheme can be devised for any antibody that eliminates or at least reduces CHO oxidizing enzyme(s) to an acceptable level.

Preparing antibodies from CHO cell cultures for conjugation

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Seattle, WA, United States of America

John Fay Valliere-Douglass