Application Notes

Studying the interaction of the antibody-drug conjugate SYD985 with an anti-toxin antibody

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1 Antibody-Drug Conjugates (ADCs) Application Note NT-MO-027 Studying the interaction of the antibody-drug conjugate SYD985 with an anti-toxin antibody Moran Jerabek-Willemsen 1 , David Egging 2 1 NanoTemper Technologies GmbH, Munich, Germany 2 Synthon Biopharmaceuticals B.V. Abstract Antibody-drug conjugates (ADCs) have been recognized as a promising new class of therapeutic agents for the treatment of cancer (1). In the generation of ADCs, cytotoxic small molecules are covalently attached to therapeutic antibodies thereby increasing their tumor cell killing capability. We have performed MicroScale Thermophoresis experiments to determine the K d values for the interaction between fluorescently labeled antibody-drug conjugate (SYD985) and an anti- toxin (drug) antibody in phosphate buffer as well in human plasma. The data show that MicroScale Thermophoresis is a powerful tool for the study of antibody-antigen interactions in standard buffers as well as complex bioliquids. Introduction SYD985 is an antibody drug conjugate composed of the monoclonal IgG1 antibody trastuzumab (SYD977) covalently bound to a linker-drug. The linker-drug contains a cleavable linker and the prodrug seco-duocarmycin-hydroxybenzamide- azaindole (seco-DUBA). The linker can be cleaved by proteases in the tumor cells at the dipeptide valine-citrulline (vc), which releases the active toxin (Figure 1). Here, we show the use of MicroScale Thermophoresis (MST) (2) to analyze the interaction of the ADC (SYD985) with a specific mouse monoclonal anti-toxin antibody. In addition we have determined the binding affinity in 50 % human plasma as well. Figure 1. Mechanism of activation of SYD985 inside the tumor cell to release the toxic duocarmycin drug. When SYD985 has been taken up into the tumor cell by endocytosis the linker is cleaved in the lysosome by proteases such as cathepsin B. Subsequently, an engineered domino-reaction occurs to generate the seco-DUBA, which then spontaneously rearranges to form the activated duocarmycin drug. This can then bind and alkylate DNA, finally resulting in cell death.

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