Emerging Concepts of Guanine Nucleotide-Binding Protein-Coupled Receptor (GPCR) Function and Implications for High Throughput Screening

Eglen, Richard M.; Bossé, Roger; Reisine, Terry

doi:10.1089/adt.2007.062

Cited by 92 publications

(78 citation statements)

References 181 publications

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“…2 and 3). This is in agreement with our previous observation that M35 and M40 behave as full agonists in a calcium flux assay performed on the same receptor (data not shown) and with the agonistic properties of the same two peptides at both GAL 1 and GAL 2 receptors. 27 We also examined NOP receptor antagonists J113397 and UFP-101.…”

Section: Discussionsupporting

confidence: 93%

G-Protein-Coupled Receptor-Mediated MAPK and PI3-Kinase Signaling Is Maintained in Chinese Hamster Ovary Cells after γ-Irradiation

et al. 2012

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To expedite G-protein-coupled receptor (GPCR) drug screening studies, cell lines amenable to transfection (e.g. CHO cells) have been widely used as cellular models. These cells can be frozen in a ready-to-use format, allowing screening of a single batch of cells and validation of the cellular material prior to the screening run. A common method used to deliver frozen cells to screening programs is to γ-irradiate the cells, abrogating cell division after thawing and ensuring consistency in the number of cells analyzed per well. With the recognition that signaling proteins such as ERK and Akt are important markers of GPCR activation, along with the availability of suitable assays for their measurement, these outputs have become important for GPCR screening programs. Here we show that several γ-irradiated and frozen CHO-K1 cell lines expressing transfected GPCRs, initially optimized for performing cAMP or AequoScreen calcium flux assays, can be used for the measurement of GPCR-mediated ERK and Akt phosphorylation. Furthermore, CHO-K1 cells transfected with NOP or GAL 1 receptors show pharmacology for a number of agonists and antagonists that is consistent with nonirradiated cultured lines. These data indicate that γ-irradiated CHO-K1 cells can be reliably used for the measurement of GPCR-mediated kinase signaling outputs.

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Section: Discussionsupporting

confidence: 93%

G-Protein-Coupled Receptor-Mediated MAPK and PI3-Kinase Signaling Is Maintained in Chinese Hamster Ovary Cells after γ-Irradiation

et al. 2012

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“…In assays with significant amplification, such as second-messenger assays (e.g., cAMP formation), both full and partial agonists can reach the same maximal response (Fig. 1A), whereas in assays with little amplification, such as assays that monitor recruitment of ␤-arrestin to a receptor by enzyme complementation (Eglen et al, 2007), partial agonists have significantly lower maximal responses than full agonists (Rajagopal et al, 2010) (Fig. 1B).…”

Section: Introductionmentioning

confidence: 99%

Quantifying Ligand Bias at Seven-Transmembrane Receptors

Rajagopal

Ahn²,

Rominger³

et al. 2011

Mol Pharmacol

361

472

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Seven transmembrane receptors (7TMRs), commonly referred to as G protein-coupled receptors, form a large part of the "druggable" genome. 7TMRs can signal through parallel pathways simultaneously, such as through heterotrimeric G proteins from different families, or, as more recently appreciated, through the multifunctional adapters, ␤-arrestins. Biased agonists, which signal with different efficacies to a receptor's multiple downstream pathways, are useful tools for deconvoluting this signaling complexity. These compounds may also be of therapeutic use because they have distinct functional and therapeutic profiles from "balanced agonists." Although some methods have been proposed to identify biased ligands, no comparison of these methods applied to the same set of data has been performed. Therefore, at this time, there are no generally accepted methods to quantify the relative bias of different ligands, making studies of biased signaling difficult. Here, we use complementary computational approaches for the quantification of ligand bias and demonstrate their application to two well known drug targets, the ␤2 adrenergic and angiotensin II type 1A receptors. The strategy outlined here allows a quantification of ligand bias and the identification of weakly biased compounds. This general method should aid in deciphering complex signaling pathways and may be useful for the development of novel biased therapeutic ligands as drugs.

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“…Identification of SM drugs with allosteric mechanisms of action came about primarily with the increasing use of functional screening assays, 96,97 but it is becoming increasingly evident that antibodies can also exhibit such modulatory effects.…”

Section: Orthosteric Versus Allosteric Mechanismsmentioning

confidence: 99%

New Horizons in Therapeutic Antibody Discovery: Opportunities and Challenges versus Small-Molecule Therapeutics

Smith¹

2015

SLAS Discovery

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Antibody drugs have become an increasingly significant component of the therapeutic landscape. Their success has been driven by some of their unique properties, in particular their very high specificity and selectivity, in contrast to the offtarget liabilities of small molecules (SMs). Antibodies can bring additional functionality to the table with their ability to interact with the immune system, and this can be further manipulated with advances in antibody engineering. This review summarizes what antibody therapeutics have achieved to date and what opportunities and challenges lie ahead. The target landscape for large molecules (LMs) versus SMs and some of the challenges for antibody drug development are discussed. Effective penetration of membrane barriers and intracellular targeting is one challenge, particularly across the highly resistant blood-brain barrier. The expanding pipeline of antibody-drug conjugates offers the potential to combine SM and LM modalities in a variety of creative ways, and antibodies also offer exciting potential to build bi-and multispecific molecules. The ability to pursue more challenging targets can also be further exploited but highlights the need for earlier screening in functional cell-based assays. I discuss how this might be addressed given the practical constraints imposed by high-throughput screening sample type and process differences in antibody primary screening.

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Emerging Concepts of Guanine Nucleotide-Binding Protein-Coupled Receptor (GPCR) Function and Implications for High Throughput Screening

Cited by 92 publications

References 181 publications

G-Protein-Coupled Receptor-Mediated MAPK and PI3-Kinase Signaling Is Maintained in Chinese Hamster Ovary Cells after γ-Irradiation

G-Protein-Coupled Receptor-Mediated MAPK and PI3-Kinase Signaling Is Maintained in Chinese Hamster Ovary Cells after γ-Irradiation

Quantifying Ligand Bias at Seven-Transmembrane Receptors

New Horizons in Therapeutic Antibody Discovery: Opportunities and Challenges versus Small-Molecule Therapeutics

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