Engineering a Potential Antagonist of Human Thyrotropin and Thyroid-stimulating Antibody

Fares, Fuad; Levi, Flonia; Reznick, Abraham Z.; Kraiem, Zaki

doi:10.1074/jbc.m008093200

Cited by 16 publications

(18 citation statements)

References 25 publications

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“…Previous attempts to dissociate binding from signal transduction have been summarized previously (70). More recent efforts using deglycosylation of linked TSH subunits resulted in engineering a potential antagonist with inhibitory effects in the in vitro bioassay (49). Certain peptides encompassing domains of the hTSH subunits show specific TSHR binding at very high concentra- tions (mM range) but do not generate a signal (130).…”

Section: Antagonistsmentioning

confidence: 99%

Thyroid-Stimulating Hormone and Thyroid-Stimulating Hormone Receptor Structure-Function Relationships

Szkudlinski

Frémont²,

Ronin³

et al. 2002

Physiological Reviews

395

256

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This review focuses on recent advances in the structure-function relationships of thyroid-stimulating hormone (TSH) and its receptor. TSH is a member of the glycoprotein hormone family constituting a subset of the cystine-knot growth factor superfamily. TSH is produced by the pituitary thyrotrophs and released to the circulation in a pulsatile manner. It stimulates thyroid functions using specific membrane TSH receptor (TSHR) that belongs to the superfamily of G protein-coupled receptors (GPCRs). New insights into the structure-function relationships of TSH permitted better understanding of the role of specific protein and carbohydrate domains in the synthesis, bioactivity, and clearance of this hormone. Recent progress in studies on TSHR as well as studies on the other GPCRs provided new clues regarding the molecular mechanisms of receptor activation. Such advances are a result of extensive site-directed mutagenesis, peptide and antibody approaches, detailed sequence analyses, and molecular modeling as well as studies on naturally occurring gain- and loss-of-function mutations. This review integrates expanding information on TSH and TSHR structure-function relationships and summarizes current concepts on ligand-dependent and -independent TSHR activation. Special emphasis has been placed on TSH domains involved in receptor recognition, constitutive activity of TSHR, new insights into the evolution of TSH bioactivity, and the development of high-affinity TSH analogs. Such structural, physiological, pathophysiological, evolutionary, and therapeutic implications of TSH-TSHR structure-function studies are frequently discussed in relation to concomitant progress made in studies on gonadotropins and their receptors.

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Section: Antagonistsmentioning

confidence: 99%

Thyroid-Stimulating Hormone and Thyroid-Stimulating Hormone Receptor Structure-Function Relationships

Szkudlinski

Frémont²,

Ronin³

et al. 2002

Physiological Reviews

395

256

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“…However, loss of glycosylation leads to the loss of TSH biological activity (27). Moreover, deglycosylated isoforms may disrupt the biological activity of normal TSH and human thyroid-stimulating immunoglobulins (28), possibly by preventing the binding of other isoforms (29,30).…”

Section: Glycosylation Patterns and Tsh Bioactivitymentioning

confidence: 99%

Thyrotropin Isoforms: Implications for Thyrotropin Analysis and Clinical Practice

Estrada

Soldin

Buckey

et al. 2014

Thyroid

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Serum thyrotropin (TSH) is considered the single most sensitive and specific measure of thyroid function in the general population owing to its negative logarithmic association with free triiodothyronine and free thyroxine concentrations. It is therefore often the test of choice for screening, diagnosis, and monitoring of primary hypothyroidism. Serum TSH concentrations can be analyzed quantitatively using third-generation immunoassays, whereas its bioactivity can be measured by TSH activity assays in cell culture. Theoretically, if serum TSH concentrations are directly related to TSH activity, the two tests should yield comparable results. However, on occasion, the results are discordant, with serum concentrations being higher than TSH biological activity. This review focuses on the dissociation between the clinical state and serum TSH concentrations and addresses clinically important aspects of TSH analysis. Thyrotropin SynthesisT hyrotropin (TSH) is a heterodimeric 28-kDa-glycoprotein hormone released from the anteromedial pituitary and is a regulator of thyroid function. Its synthesis is controlled by the hypothalamic neuropeptide TSH-releasing hormone (TRH). The two peptide subunits of TSH are noncovalently linked and cotranslationally glycosylated with mannose-rich oligosaccharides (1). Posttranslationally, the two subunits are combined and the attached oligosaccharides are further processed. Synthesis of a mature TSH molecule requires the excision of signal peptides from both TSH a-and b-subunits, followed by trimming of mannose and further addition of fucose, galactose, and sialic acids (2). Thus, mature TSH molecules are asparagine-(N)-linked [Asp(N)-linked] complex carbohydrate structures capped with sulfate and/or sialic acid molecules (3,4) (Fig. 1). TSH oligosaccharide structures vary according to the source of TSH: human pituitaryderived TSH comprises fucosylated biantennary glycans with terminal N-acetylgalactosamine sulfate and low sialic acid content (5-7), while recombinant human TSH (rhTSH), synthesized in Chinese hamster ovary cells, specifically terminates in a2,3-linked sialic acid (8-11) and rhTSH formed in yeast lacks sialic acid residues altogether (12). Glycosylation Patterns and TSH BioactivityAppropriate glycosylation is necessary to maintain normal TSH bioactivity (13). Indeed, both a-and b-subunits of TSH have functionally important domains associated with TSH receptor (TSHR) binding and activation. The a-subunit has two Asp(N)-linked oligosaccharide chains with a typical biantennary structure, while TSH b-subunit has only one chain (Fig. 2). The transcriptional and posttranscriptional mechanisms involved in TSH glycosylation result in folding of these subunits, leading to their heterodimerization. These mechanisms also qualitatively regulate TSH secretion, prevent intracellular degradation, and influence TSH clearance rate from the circulation (14,15). A three-dimensional TSH structure has been proposed in the 1990s based on crystallographic studies and comparisons with other glyco...

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“…To autonomously control thyroid hormone homeostasis in a selfsufficient manner, we coupled the TSR sensor device with the expression of an engineered high-affinity TSH Antag that competes with endogenous TSH for the TSHR-binding site (25). TSH Antag is an engineered variant of human TSH containing a human chorionic gonadotrophin-β C-terminal peptide linker between the β-and α-chains as well as two mutated glycosylation sites (18).…”

Section: Closed-loop Control Of Tsh Antag Secretion By the Thyroid Homentioning

confidence: 99%

Synthetic gene network restoring endogenous pituitary–thyroid feedback control in experimental Graves’ disease

Saxena

Hamri

Folcher

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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Graves’ disease is an autoimmune disorder that causes hyperthyroidism because of autoantibodies that bind to the thyroid-stimulating hormone receptor (TSHR) on the thyroid gland, triggering thyroid hormone release. The physiological control of thyroid hormone homeostasis by the feedback loops involving the hypothalamus–pituitary–thyroid axis is disrupted by these stimulating autoantibodies. To reset the endogenous thyrotrophic feedback control, we designed a synthetic mammalian gene circuit that maintains thyroid hormone homeostasis by monitoring thyroid hormone levels and coordinating the expression of a thyroid-stimulating hormone receptor antagonist (TSHAntag), which competitively inhibits the binding of thyroid-stimulating hormone or the human autoantibody to TSHR. This synthetic control device consists of a synthetic thyroid-sensing receptor (TSR), a yeast Gal4 protein/human thyroid receptor-α fusion, which reversibly triggers expression of the TSHAntag gene from TSR-dependent promoters. In hyperthyroid mice, this synthetic circuit sensed pathological thyroid hormone levels and restored the thyrotrophic feedback control of the hypothalamus–pituitary–thyroid axis to euthyroid hormone levels. Therapeutic plug and play gene circuits that restore physiological feedback control in metabolic disorders foster advanced gene- and cell-based therapies.

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Engineering a Potential Antagonist of Human Thyrotropin and Thyroid-stimulating Antibody

Cited by 16 publications

References 25 publications

Thyroid-Stimulating Hormone and Thyroid-Stimulating Hormone Receptor Structure-Function Relationships

Thyroid-Stimulating Hormone and Thyroid-Stimulating Hormone Receptor Structure-Function Relationships

Thyrotropin Isoforms: Implications for Thyrotropin Analysis and Clinical Practice

Synthetic gene network restoring endogenous pituitary–thyroid feedback control in experimental Graves’ disease

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