G protein-coupled receptors (GPCRs) regulate nearly every aspect of human physiology and diseases, and form one of the largest drug target families. GPCRs share a conserved structure with 7 transmembrane α-helices. Antibodies binding to a GPCR protein on the cell surface can be a very useful tool for target detection and potential drug discovery. The structure complexity and dynamic conformations make it very challenging to develop antibodies binding GPCRs on live cell surfaces. To advance GPCR antibody discovery, we have developed an innovative fit-for-target precision antibody discovery platform to generate GPCR-specific monoclonal antibodies (mAbs).
GPCR protein sequence and structure for each target was analyzed using multiple parameters. Based on each target’s unique sequence and structure features, we design fit-for-target immunogens, formulations and immunization regimens and conduct immunizations in mice. Immunized mouse sera were evaluated for antibody responses against each specific GPCR target antigen by ELISA and target expressing cells by flow cytometry. Then, immune mouse B cells were isolated to generate target-specific mAbs using hybridoma technology. The GPCR target-specific mAbs were characterized for binding specificity and live cell staining using multiple assays including ELISA and flow cytometry.
We selected a large panel of GPCR targets from Class A, B, C and F to generate extracellular domains specific antibodies. High titers of target-specific antibody responses were elicited in mice. The immune mouse sera could bind to the relevant specific target antigen by ELISA and target expressed on live cell surfaces by flow cytometry. The GPCR target-specific mAbs were then generated using hybridoma technology. Most of these mAbs had strong binding affinity and excellent specificity against the relevant GPCR antigen as measured by ELISA and SPR. Many of the mAbs could bind to GPCR expressed on live cell surfaces measured by flow cytometry. These mAbs target a panel of GPCRs related to major therapeutic areas, including immune & inflammation (ADGRE2, CRFR2, GPR183, CX3CR1), neurology (GPR52, CALRL, DRD2), cardiovascular (APLNR, PAR1, MAS1, EDNRB), metabolic (GLP1R, GCGR, GIPR), and oncology (LGR4, GPR56, FZD10).
Conigen’s innovative fit-for-target precision antibody discovery platform can effectively generate monoclonal antibodies against challenging GPCRs. The novel GPGR-specific mAbs are very useful tools for research and drug discovery.
