Calcitonin Gene-Related Peptides

Calcitonin-gene related peptide (CGRP), amylin (AMY), adrenomedullin (AM), calcitonin receptor-stimulating peptide (CRSP) and intermedin/adrenomedullin-2 (IMD/AM-2) are structurally related peptides of the same family. They are characterized by a six to seven amino acid ring structure linked by a disulfide bridge and an amidated C-terminus.

CGRP is a 37 amino acid residue peptide derived from the alternative splicing, in a tissue specific manner, of the calcitonin gene and may exist in two forms α and β with similar biological functions. CGRP is widely distributed in both the central and peripheral nervous systems and exhibits of range of biological actions. Structure-activity studies suggest that these effects are mediated by the activation of at least two CGRP receptor subtypes designated as CGRP₁ and CGRP₂. This classification is based on the preferential affinity of the fragment CGRP(8-37) to antagonize the effect of CGRP (CGRP₁), and the agonistic property of the linear analog [Cys(ACM)^2,7]hCGRPa to mimic the effect of CGRP (CGRP₂). The use of BIBN4096BS, the first non-peptide CGRP antagonist having sub-nanomolar affinity for CGRP receptors, has led to the discovery of further functional CGRP receptor heterogeneity in rodent tissues. BIBN4096BS was much less potent at antagonizing the effects of CGRP proposed to be mediated by the CGRP₂ subtype. An orphan receptor originally described as the calcitonin receptor-like receptor (CLR) has been identified as a CGRP receptor. Its association with the single membrane-spanning receptor-activity-modifying protein 1 (RAMP1) is required for this receptor to behave as a CGRP₁ receptor. It also requires the presence of the receptor component protein (RCP) to generate a complete CGRP’s response.

AMY is a 37 amino acid peptide isolated from amyloid deposits of human insulinoma and from the pancreas of type 2 diabetic patients. AMY-like immunoreactivity is detected in peripheral tissues as well as in hypothalamic areas of the rat brain. In the CNS, AMY induces an anorexic effect similar to that of CGRP. [¹²⁵I]-AMY binding sites have been detected in the rat brain and this distribution profile is distinct from that of [¹²⁵]-CGRP sites suggesting the existence of unique amylin receptors. AC187, a salmon calcitonin derivative, behaves as a competitive amylin receptor antagonist. At the molecular level, it appears that the calcitonin receptor co-transfected with RAMPs generates three amylin receptor subtypes.

AM is a polypeptide of 50 amino acids in length in the rat and of 52 amino acids in human. AM-like immunoreactivity and corresponding mRNA are present in various rat and human tissues. Administration of AM produces biological effects similar to those reported for CGRP. It was thus proposed that these two peptides were acting via a common CGRP-like receptor since the potent vasodilation action and anorexic effect of AM were antagonized by CGRP(8-37). However, several studies have demonstrated that compared with AM, CGRP is 100- to 1000-times less potent at competing for [¹²⁵I]-AM(1-52) and [¹²⁵I]-AM(13-52) binding sites in various assays. Additionally, structure-activity studies revealed that removing the first 21 N-terminal amino acid residues of hAM, just next to the disulfide bridge (hAM(22-52)), resulted in a peptide with antagonist properties. CLR appears to function as an AM receptor when co-expressed with RAMP2. The existence of additional AM receptor subtypes is supported by functional assays using cells cotransfected with CLR and RAMPs.

Other peptides have been identified and been added to this peptide family. Three porcine CRSPs are related to CGRP but the biological effects of CRSP-1 are mediated through the calcitonin receptor. CRSP-like immunoreactivity and mRNA have mainly detected in the brain, pituitary and thyroid gland. CRSPs were also identified in dog, cow and horse but not in human and rodents. IMD/AM-2, a 47 amino acid peptide related to AM, is the most recently discovered member of this peptide family. IMD/AM-2-like immunoreactivity and corresponding mRNA are expressed in various tissues including the submaxillary gland, kidney, lung, heart, gastrointestinal system, pituitary gland and hypothalamus, but not in the adrenal gland. Recent studies have demonstrated that IMD/AM-2 acts on CLR/RAMP-like receptors and exerts biological effects similar to those reported for CGRP and AM, such as cardiovascular regulation and decreased food and water intake.

The Table below contains accepted modulators and additional information. For a list of additional products, see the "Related Products" section below.

Currently Accepted Name	CGRP1	CGRP2
Cloned Receptor and Associated Proteins	CLR (464 aa) RAMP1 (148 aa) RCP (146 aa)	Not Known
Potency of Endogenous Homologs	CGRPα (C0167) ~ CGRPβ > IMD/AM-2 = AM (A3830) ≥ AMY > CT	CGRPα (C0167) ~ CGRPβ > AM (A3830) > AMY
Preferential Agonist	Not Known	[Cys(ACM)2,7]-hCGRP
Antagonists	BIBN4096BS CGRP(8-37) (C2806) SB-273779 (SML0391)	BIBN4096BS CGRP(8-37) (C2806)
Prototypical Tissue Bioassays	Atrium Pulmonary artery Spleen SK-N-MC cells	Vas deferens Urinary bladder Liver Col-29 cells HCA-7 cells
Signal Transduction Mechanisms	Gs (cAMP production)	Gs (cAMP production)
Radioligands of Choice	[125I]-[Iodohistidyl10]-hCGRPα [125I]-hCGRP(8-37)	[125I]-[Iodohistidyl10]-hCGRPα
Tissue Expression	CNS PNS Cardiovascular tissues Blood vessels Spleen Vas deferens Lung Adrenal gland	Coronary arteries Vas deferens Urinary bladder Liver
Physiological Function	Regulation of blood pressure, heart rate Respiratory regulation Calcium homeostasis Control of fluid balance, diuresis, natriuresis Inhibition of gastric acid secretion Inflammation Nociception	Not Known
Disease Relevance	Migraine Hypertension Pulmonary hypertension Septic shock	Not Known

Currently Accepted Name	AM1	AM2	AMY
Cloned Receptor and Associated Proteins	CLR RAMP2 (182 aa)	CLR RAMP3 (148 aa)	CTR (516 aa) + RAMP1 known as AMY1 + RAMP2 known as AMY2 + RAMP3 known as AMY3
Potency of Endogenous Homologs	AM (A3830) > IMD/AM-2 = CGRPα (C0167)  ~ CGRPβ ≥ AMY > CT	AM (A3830) > IMD/AM-2 ≥ CGRPβ > CGRPα (C0167) ≥ AMY > CT	sCT (T3660) ~ rAMY >> hCT (T3535) CGRP and AM ligand selectivity remains to be fully established.
Preferential Agonist	[Mpr14]-rAM(14-50)	Not Known	AMY
Antagonists	AM(22-52) > CGRP(8-37) (C2806)	CGRP(8-37) (C2806) > AM(22-52)	AC187 AC253 (SML0625) AC413 AC512 sCT(8-32) The antagonist preference for each AMY subtype is still to be established.
Prototypical Tissue Bioassays	Vascular smooth muscle, lung	Not Known	Skeletal muscle, MCF-7 cells, ovary cell line CHO-K1
Signal Transduction Mechanisms	Gs (cAMP production)	Gs (cAMP production)	Gs (cAMP production)
Radioligands of Choice	[125I]-AM(1-52) [125I]-AM(13-52)	[125I]-AM(1-52)	[125I]-Amylin [125I]-Salmon calcitonin
Tissue Expression	CNS PNS Cardiovascular tissues Spleen Vas deferens Lung Liver Skeletal muscle	Not Known	CNS Lung Stomach Spleen Liver Kidney
Physiological Function	Regulation: blood pressure (vasorelaxation) heart rate Control of: fluid balance diuresis natriuresis angiogenesis cell growth Regulation of food intake Nociception	Not Known	Reduction of food intake Glucose regulation Inhibition of gastric emptying and acid secretion Inhibition of pancreatic enzymes Calcium homeostasis Memory retention
Disease Relevance	Hypertension Congestive heart failure Pulmonary hypertension Septic shock Renal failure Cerebral ischemia	Not Known	Diabetes Anoxeria

Abbreviations

ACM: Acetamidomethyl
AC187: Acetyl-[Asn³⁰,Tyr³²] salmon calcitonin (8–32)
AM: Adrenomedullin
AMY: Amylin
BH: Bolton Hunter labeled
BIBN4096BS: [r-(r*,s*)]-N-[2-[[5-Amino-1-[[4-(4-pyridinyl)-1-piperazinyl]carbonyl]pentyl]amino]-1- [(3,5-dibromo-4-hydroxyphenyl)methyl]-2-oxoethyl]-4-(1,4-dihydro-2-oxo-3(2h)-quinazolinyl)-,1-piperidinecarboxamide
CGRP: Calcitonin gene-related peptide
CLR: Calcitonin receptor-like receptor
CNS: Central nervous system
CTR: Calcitonin receptor
hCT: Human calcitonin
IMD/AM-2: Intermedin/Adrenomedullin-2
PNS: Peripheral nervous system
RAMP: Receptor-activity-modifying protein
RCP: Receptor component protein
SB-273779: N-Methyl-N-(2-methylphenyl)-3-nitro-4-(2-thiazolylsulfinyl)nitrobenzanilide
sCT: Salmon calcitonin

h: human
s: salmon

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