G protein-coupled receptors (GPCRs) represent a major therapeutic target class as they play a key role in many (patho)physiological processes. Besides small molecules, GPCRs can be effectively targeted by biologics. In particular, antibody fragments from camelid-derived heavy chain-only antibodies (nanobodies/VHHs) are attractive tools in detecting, stabilizing, modulating and therapeutically targeting GPCRs. The small size and molecular structure of nanobodies allow extracellular and intracellular modulation of GPCR function. Besides modulating GPCR activity as monovalent or multivalent constructs, nanobodies can also be functionalized for imaging and therapy. Examples of nanobodies targeting human and viral chemokine receptors will be provided. Moreover, GPCR-binding nanobodies have been instrumental in obtaining crystal structures of GPCRs, facilitating structure-based drug discovery. Thus, nanobodies are an important class of biologics targeting GPCRs. 

Despite considerable interest in antibody-based therapeutics targeting GPCRs, few reported antibodies functionally modulate GPCR signaling. We developed a methodology to discover modulatory camelid antibody fragments (nanobodies) for GPCRs. Using this strategy, we identified nanobody antagonists for the angiotensin II type I receptor (AT1R), which regulates cardiovascular homeostasis. Our nanobody performs similarly to clinically used angiotensin receptor blockers (ARBs) and provides a strategy for targeting AT1R when ARBs are contraindicated.

Developing functional antibodies targeting G-protein-coupled receptors remains challenging. Here we report structure-based and function-based approaches for generating functional single domain antibodies (sdAbs) against human apelin receptor (huAPJ). We co-crystallized an orthosteric sdAb antagonist complexed with huAPJ and converted it into an agonist by structure-guided design. We further developed an HTS method for directly isolating functional antibodies against GPCRs and identified a panel of sdAb antagonists and an agonist to huAPJ.

Human cell microarray screening enables the discovery of specific primary cell surface receptors (i.e., druggable targets) as well as uncovering off-targets for a variety of biotherapeutic molecules, including peptides, antibodies and proteins, CAR T and other cell therapies. Case studies will focus on the assessment of off-target liabilities at an early stage and highlight the increasing role of the technology as an adjunct to, or replacement for, tissue cross reactivity studies prior to IND submissions.

G protein-coupled receptors (GPCRs) and ion channels represent some of the most important membrane protein drug target classes across a wide range of therapeutic areas. An update on antibody-based therapeutics in the GPCR/ion channel pipeline will be provided outlining the breadth and diversity of the target landscape, progress in preclinical and clinical development, including examples that highlight the successes and challenges faced with these target classes.

The NRC aims to advance a functionally relevant conformational GPCR antigen presentation technology for high efficiency generation of functional mAbs for various therapeutic indications. Considering multiple examples, we have demonstrated that some mAbs identified behave as PAM or NAM (positive or negative allosteric modulators). This presentation will demonstrate the value of our GPCR platforms to identify lead candidates that target diverse intrinsic pharmacological properties selective toward active or inactive GPCR conformations.

G-protein coupled receptors play key roles in sensorial, metabolic, immunological and neurotransmission processes.  Despite its importance, quantifying the binding of drug candidates are difficult to do with traditional SPR techniques.  Results of kinetic measurements using SPR microscopy on GPCR and certain small molecules and antibodies will be shown in detail.

The voltage-gated sodium channel Nav1.7 is critical for amplifying pain signals. Selective blockers of Nav1.7 could provide analgesia with minimal side effects. One source of Nav1.7 drug discovery starting points are spider venoms that block voltage-gated sodium channels. This talk will discuss engineering efforts on huwentoxin-IV and protoxin-II, alterations in them that produced favorable changes in Nav1.7 potency and/or selectivity, and their ability to pharmacologically recapitulate the Nav1.7-null phenotype.

We have identified various agonists of TRPA1 channel, and determined their ligand-binding and gating mechanism through biophysical and structural studies. Different from electrophilic agonists, non-covalent agonists activate TRPA1 without desensitization or tachyphylaxis, and evoke pain responses with different kinetics and sensitivity to antagonist treatment. Our study provides a pathway for the discovery of drugs tailored to specific disease conditions.

Transporter proteins control the movement of nutrients and other molecules across the cell membrane and are targets for diseases including diabetes and cancer. The structural complexity of transporters including SLC2A4 (GLUT4) pose a barrier in discovering MAbs for research and therapeutics. We leveraged our MPS Antibody Discovery platform to discover the first functional, conformation-dependent MAbs against SLC2A4. These MAbs selectively bind state-specific active forms of the transporter, providing new molecular tools for therapeutic discovery.

Resistance to antimalarials such as chloroquine (CQ) and piperaquine (PPQ) has been associated with distinct sets of point mutations in the P. falciparum chloroquine resistance transporter PfCRT, a 49 kDa integral transmembrane protein localized in the digestive vacuole of the pathogenic parasite. We present the 3.2 Å structure of a PfCRT isoform from CQ-resistant, PPQ-sensitive South American 7G8 parasites, using a combination of single-particle cryo-electron microscopy and fragment antigen-binding technology.

Twist Biopharma provides antibody discovery and optimization solutions.  To discover functional GPCR antibodies, we grafted GPCR-binding motifs into an antibody library.  Another approach mined the patented GPCR antibody sequences and used them to design another GPCR-focused library.  We can discover potent functional antibodies against GPCR targets using both libraries.