Protease-activated (Thrombin) Family Related Drug Discovery Products

Protease-activated receptors (PARs) belong to the family of GPCRs. Proteases specifically cleave the amino terminal sequence of these receptors, exposing a new N-terminal region that functions as a tethered ligand. This ligand then binds intramolecularly to activate the receptor. Although they are found everywhere in the body, the neurological system, vascular, immunological, and intestinal epithelial cells express PARs most frequently. As a result, a number of vital biological processes involved in physiology and illness are controlled by their activations.

Fig.1. Mechanisms of protease-activated receptors activation. (Sébert, 2019)

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Overview of Protease-activated (Thrombin) Family

• F2R (PAR₁)

By producing RNA from thrombin-responsive cells from humans and hamsters in Xenopus oocytes, the gene for PAR₁ was successfully cloned. Clones that encoded a 425 residue protein with 7 typical hydrophobic GPCR domains were discovered. Human PAR₁'s inferred sequence included an amino-terminal signal sequence, a 75-residue extracellular amino-terminal domain, and a putative thrombin cleavage site in the amino-tail.

• F2RL1 (PAR₂)

Using degenerate primers targeting the second and sixth transmembrane domains of the bovine neurokinin 2 receptor, PAR₂ was discovered by screening a mouse genomic library. A 395 residue protein with typical GPCR properties and ~30% amino acid identity to human PAR₁ was identified to be encoded by a clone. A potential trypsin cleavage site was present at the amino terminus of 46 residues that was extracellular.

• F2RL2 (PAR₃)

The discovery that platelets from PAR₁-deficient mice responded to thrombin whereas fibroblasts did not suggested the existence of additional receptors for thrombin. In order to filter RNA from rat platelets, PAR₃ was subsequently cloned using degenerate primers to conserved regions of PAR₁ and PAR₂. After being cloned in both human and murine forms, it was discovered that human PAR₃ has ~28% of its sequence homology with human PAR₁ and PAR₂. Similar to PAR₁ and PAR₂, PAR₃ is a conventional GPCR with an extracellular amino terminus thrombin cleavage site.

• F2RL3 (PAR₄)

Human PAR₄ is a 385-amino acid protein. It may be cleaved by thrombin and trypsin at its extracellular amino-terminal domain. Although PAR₄ and the other human PARs share 33% of their amino- and carboxy-terminal domains, there are some noticeable distinctions.

Fig.2. Cleavage site of PARs. (Han, 2021)

Protease-activated (Thrombin) Family Drug Discovery

It has been established that PARs have a significant role in the control of the physiological functions of the intestine as well as the two major illnesses that affect the organ, inflammatory bowel disorders and colorectal cancer. As a result, the creation of therapeutic drugs is attracted to these G-protein coupled receptors. Though effective PAR antagonist development is still in its early stages, attempts to develop selective receptor inhibitors are now prioritized over targeting their ligands.

References

1. Sébert, M.; et al. Protease-activated receptors in the intestine: focus on inflammation and cancer. Frontiers in Endocrinology. 2019, 10: 717.
2. Han, X.; et al. Protease‐activated receptors: An illustrated review. Research and Practice in Thrombosis and Haemostasis. 2021, 5(1): 17-26.