The Thyrotropin receptor (TSHR) is a central regulator of growth and metabolism. Euthyroid states are marked by synthesis and secretion of thyroid hormones upon TSHR activation. However, aberrant TSHR activity plays a major etiological role in autoimmune thyroid diseases. How TSHR autoantibodies modulate receptor activity has remained elusive. Using cryo-EM, molecular dynamics, and signaling assays, we determined principles of TSHR activation and a hormone-mimicry mechanism employed by a patient-derived autoantibody. Our results suggest that ligands interacting with the TSHR extracellular domain (ECD) elicit their activity through steric constraint of ECD orientations.

This study aims to understand conformational states of the coreceptor CCR5. Based on CCR5 conformational changes, down-modulation, and trafficking rates, we were able to distinguish which among heterogeneous CCR5 subpopulations might best be targeted to inhibit HIV-1 entry. We assume that a decreased surface presence of a particular CCR5 subpopulation following infection means that it has been internalized due to HIV-1 entry and that it, therefore, represents a highly relevant target for antiretroviral therapies. Our work may lead to the future identification of better HIV-1 entry inhibitors of prophylactic or therapeutic efficacy and may contribute to future HIV-1 vaccine designs. 

ConfoBodies, single-domain antibody fragments from camelids (VHH) stabilize a desired conformational state of a GPCR and enable conformation-directed drug screening. We will show the unique potential of ConfoBody-stabilized GPCR conformations to facilitate de novo discovery of therapeutic antibodies exhibiting full agonist pharmacology to human GPCRs.

The GPCRs for lipid mediators such as prostaglandins and lysophosphatidic acid are promising targets for various diseases. The generation of therapeutic mAb targeting GPCR for lipid mediators is more difficult than mAb for chemokine and peptide GPCRs. NBHL established MoGRAA discovery engine which unlocks the generation of mAbs targeting lipid mediator GPCR. MoGRAA discovery engine includes proprietary DNA immunization and a unique single-cell analysis. In addition, the various profiling including functional assay, specificity test, and phenotypic assay are crucial steps. We will introduce mAbs targeting Prostaglandin E2 and lysophosphatidic acid receptors for cancer and fibrosis as case studies.

GPCR agonist antibody discovery is challenging due to the complexities of receptor activation. Leveraging partners, we employed non-traditional discovery approaches to create an agonist antibody. Chicken immunization for antibody diversity provided an interaction capable of activating a GPCR that no other rodent or human phage display derived antibodies could in the same epitope region—delivering an agonist mAb with robust pharmacodynamic activity, good pharmacokinetics, and minimal pre-development sequence liabilities.

The NALCN leak channel regulates the resting membrane potential of many neurons. NALCN modulates locomotion, circadian rhythm and respiration, and mutations in NALCN cause severe neuro-developmental disorders. We determined the structure of the NALCN channelosome, an approximately 1-MDa complex consisting of NALCN and four auxiliary subunits. Our findings provide a blueprint to understand the physiology of NALCN and a foundation for drug discovery to treat NALCN channelopathies.

Membrane proteins are important drug targets (GPCRs, ion channels), yet are notoriously difficult to work with. We’ll present the direct extraction of ion channels from crude cells as well as case studies on Salipro-embedded ion channels (e.g. TRPV3) for phage display, immunization, B cell sorting, and antibody characterization with SPR and high-resolution cryoEM.

Transient receptor potential ankyrin 1 (TRPA1), a nonselective calcium channel highly expressed in primary sensory neurons and involved in the induction of airway inflammation and hyperreactivity, is a target for inhibition in asthma. Our structural studies using cryoEM have unveiled the binding sites and mechanisms of action of two distinct chemical series of antagonists. In addition, we characterized a non-covalent agonist and found that it elicits pain responses that are distinct from those of previously described agonists, correlating with differentiated conducting states and desensitization.

Voltage-gated sodium channels are responsible for the rapid rise of action potentials in excitable tissues, and NaVchannel mutations have been implicated in several human genetic diseases. We generated high-affinity anti-NaV nanobodies that recognize the NaV1.4 and NaV1.5 channel isoforms. Our work lays the foundation for developing Nbs as anti-NaV reagents to capture NaVs from cell lysates and as molecular visualization agents for NaVs.

Prolonged glycine signaling has long been a therapeutic goal to improve cognitive impairment and the negative symptoms in patients with schizophrenia. Partially effective, it remained unclear how inhibitors of the glycine transporter 1 (GlyT1) bind to restore normal levels of glycine. Using structure biology, we create an understanding of the binding mode of a benzoylpiperazine chemotype inhibitor bound to GlyT1 and enable the rational design of novel chemotypes.

Docosahexaenoic acid is an essential omega-3 fatty acid that is transported across the blood-brain barrier by MFSD2A. Here we solved the cryo-EM structure of MFSD2A in complex with an Fab and lysolipid substrate. Together with functional assays and molecular dynamics simulations, this structure reveals details of how MFSD2A interacts with substrates and releases them into the membrane through a lateral gate. These findings have the potential to aid the delivery of neurotherapeutics into the brain.

The ability to detect and characterize drug binding to SLC transporters is of growing interest in academic and industrial drug discovery labs. However, SLC purification and binding characterization to SLC targets of interest are extremely challenging; therefore, very few targeted discovery programs have been able to move forward into meaningful preclinical studies in this particular space. Here, we selected MCT1, an SLC target of interest in the oncology field, as a case study to demonstrate initial proof-of-principle to perform label-free measurement of target engagement to integral MCT1 in its native cell membrane environment. This work emerged from a collaboration between Amgen and Nanotech Bio.