Characterising therapeutic antibodies and ADCs using mass spectrometry

24 August 2016  •  Author(s): Ioannis Papayannopoulos, Celldex Therapeutics

Since the 1970s, the advent of biotechnology has resulted in the development and commercialisation of many therapeutic proteins, including antibodies and antibody fragments, for the treatment of human diseases. Examples include antibody treatments for autoimmune diseases (for example, adalimumab [Humira®] for rheumatoid arthritis), cancers (such as trastuzumab [Herceptin®] for breast cancer), degenerative conditions (including ranibizumab [Lucentis®] for macular degeneration), viral and bacterial infections, as well as others. Additionally, antibodies are used as imaging reagents, linked to radionuclides or non-radioactive rare earth metals, and quite extensively employed as reagents in biomedical research, commercial medical tests and clinical laboratory assays (for example, enzyme-linked immunosorbent assay [ELISA]).

Characterising therapeutic antibodies and ADCs using mass spectrometry

Therapeutic antibodies are typically monoclonal antibodies (MABs) of the immunoglobulin G (IgG) isotype, with affinities against specific antigens and which bind monospecifically to particular cells or proteins. They work either by stimulating the patient’s immune system, thus priming or enhancing the immune response, or by binding to specific molecules and thus modulating or inhibiting certain biochemical pathways involved in disease. Monoclonal antibodies have also been developed with affinities towards specific tumour antigens to deliver radionuclides to them, thereby killing tumour cells with minimal deleterious effects on non-tumour cells. Such radiommunotherapeutic antibodies are modified with a chelator that binds a radioactive metal ion that is thus transported to the tumour.


Figure 1: Schematic representation of an antibody. Enzymatic cleavage at the hinge region results in the generation of heavy and light chains that are linked via a disulfide bond (Fab). Enzymatic cleavage below the disulfide bonds that link the heavy chain constant regions 2 results in the generation of a fragment consisting of a pair of disulfide-linked heavy and light chains that are also linked to one another (F(ab’)2)

Antibody-drug conjugates (ADCs), used in cancer chemotherapy, have small-molecule chemotherapeutics covalently attached via a linker that is cleaved at the tumour site, thereby releasing a cytotoxic small molecule in the immediate vicinity of a tumour. Although only a few ADCs have received regulatory approval so far, they comprise a very active area of research and development in many biopharmaceutical companies.

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