Engineering antibody-drug conjugates targeting an adhesion GPCR, CD97

Area of Science:

• Cell Biology
• Immunology
• Oncology

Background:

• Adhesion G protein-coupled receptors (aGPCRs) are crucial cell-adhesion molecules implicated in various cellular functions and human diseases, notably cancer.
• While aGPCRs possess large extracellular regions suitable as antigens, their therapeutic potential as targets for biologic drugs remains underexplored.
• CD97 (ADGRE5) is an aGPCR frequently upregulated in cancers such as acute myeloid leukemia (AML) and glioblastoma (GBM), including their associated cancer stem cells.

Purpose of the Study:

• To develop and evaluate antibody-drug conjugates (ADCs) targeting the cell-adhesion molecule CD97 for potential therapeutic applications against AML and GBM.
• To identify a CD97-specific antibody with efficient internalization properties and assess its binding characteristics.
• To investigate the therapeutic efficacy and safety profile of CD97-targeting ADCs in preclinical cancer models.

Main Methods:

• Generation of a panel of synthetic human antibodies targeting distinct CD97 epitopes.
• Characterization of antibody binding specificity, including cross-reactivity with homologs like EMR2, and internalization efficiency.
• Structure determination of the antibody-target interaction using single-particle cryo-electron microscopy.
• Screening of conjugation strategies and cytotoxic payloads to create stable and effective ADCs.
• Assessment of ADC efficacy against AML and GBM cell lines and patient-derived GBM stem cells, alongside cytotoxicity evaluation in peripheral blood mononuclear cells.

Main Results:

• An antibody targeting the conserved GPCR autoproteolysis-inducing (GAIN) domain of CD97 was identified, exhibiting efficient internalization and specific binding across all CD97 isoforms without cross-reactivity to EMR2.
• The antibody demonstrated an unconventional binding mode, extensively utilizing its light chain framework for antigen recognition.
• A stable ADC was successfully developed, demonstrating potent killing of AML and GBM cell lines, as well as patient-derived GBM stem cells.
• The developed ADC exhibited minimal cytotoxicity against healthy donor peripheral blood mononuclear cells, indicating a favorable safety profile.

Conclusions:

• CD97 represents a promising therapeutic target for cancers like AML and GBM.
• The aGPCR GAIN domain is a viable target for antibody-based therapeutics.
• The study highlights an uncharacterized antibody binding surface on the aGPCR GAIN domain, expanding the scope of antibody recognition strategies.
• The developed CD97-targeting ADCs show significant therapeutic potential with a promising safety margin for treating specific hematologic and brain cancers.