US 9,958,455 B2 · Intact mass determination of protein conjugated agent compounds

Abstract

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.

Claims34

Dependent claims

A method for quantitating a distribution of antibody drug conjugate compounds, in a mixture comprising:

(a) providing the mixture in a matrix to a separation media run under non-denaturing conditions with a mobile phase to effect removal of the matrix from the mixture and produce an eluate;
(b) introducing the eluate into a mass spectrometer;
(c) directly establishing a mass and an ion intensity for each species of the antibody drug conjugate compounds from a deconvoluted mass spectrum;
and
(d) quantitating the distribution of the antibody drug conjugate compounds by ion intensity;
wherein:
each species of the antibody drug conjugate compounds has from one to eight drug moieties conjugated to reduced interchain disulfides, is non-covalently associated and non-denatured, and maintains its intact-bivalent antibody structure, and the mobile phase comprises a volatile salt, is free of non-volatile salts, and is compatible for use with mass spectrometry.

4 more elements

Dependencies are inferred from the claim text.

Intact mass determination of protein conjugated agent compounds

Life of this patent

Intact mass determination of protein conjugated agent compounds

Life of this patent

Abstract

Claims34