cgrp as the target of new migraine therapies — successful ... · of medications that are...

Patients with migraine — a chronic primary headache disorder that afflicts almost 15% of the population worldwide1 — will soon have access to a novel group of medications that are effective in treating or prevent-ing the debilitating, painful headaches associated with the disorder. These new therapies use a unique strategy to treat migraine, and work by interfering with the sig-nalling of a potent neuropeptide expressed in sensory nerves: calcitonin gene- related peptide (CGRP).

Migraine is a chronic, complex neurological disorder that manifests as recurrent attacks of moderate to severe headache pain lasting 4–72 h. The headache is typically unilateral, has a pulsating quality, is aggravated by rou-tine physical activity and is associated with nausea and/or sensitivity to light and sound2. In migraine with aura, the headache phase is preceded by reversible focal neu-rological symptoms, often visual or sensory, that usually develop gradually over 5–20 min and last for <60 min. People who experience frequent migraine attacks are con-sidered to have either episodic migraine (1–14 migraine days per month) or chronic migraine (>15 migraine days per month).

Previous treatment options have not met the chal-lenge of alleviating the personal and economic burden

of migraine, which is the leading cause of neurological disability and one of the top five causes of all long- term disability1,3. The success of older and current drugs for migraine prevention and abortion has been limited by inadequate efficacy, tolerability and patient adherence4–6. Drugs that are currently taken for prophylaxis are not specific for migraine but include antiepileptic drugs, β- blockers and antidepressants7,8.

All CGRP- targeted therapies tested for the treatment of migraine to date have consistently produced positive results9,10 (Table 1). CGRP receptor antagonists pro-vide relief from an acute headache attack, either with or without aura. Monoclonal antibodies against either CGRP or its receptor are particularly effective for the prevention of episodic or chronic migraine. The drugs currently being developed are well tolerated and have few adverse effects. Thus, identification of CGRP as a specific therapeutic target for migraine has resulted in a major breakthrough in providing relief for this com-mon disorder. The translation of CGRP therapies to the clinic has been a long journey of discovery (Fig. 1). As this journey comes to fruition, in this Review, we sum-marize what has been learned about the role of CGRP in migraine and the trigeminovascular pain pathway and

CGRP as the target of new migraine therapies — successful translation from bench to clinicLars Edvinsson 1,2*, Kristian Agmund Haanes2, Karin Warfvinge1,2 and Diana N. Krause1,3

Abstract | Treatment of migraine is on the cusp of a new era with the development of drugs that target the trigeminal sensory neuropeptide calcitonin gene- related peptide (CGRP) or its receptor. Several of these drugs are expected to receive approval for use in migraine headache in 2018 and 2019. CGRP- related therapies offer considerable improvements over existing drugs as they are the first to be designed specifically to act on the trigeminal pain system, they are more specific and they seem to have few or no adverse effects. CGRP receptor antagonists such as ubrogepant are effective for acute relief of migraine headache, whereas monoclonal antibodies against CGRP (eptinezumab, fremanezumab and galcanezumab) or the CGRP receptor (erenumab) effectively prevent migraine attacks. As these drugs come into clinical use, we provide an overview of knowledge that has led to successful development of these drugs. We describe the biology of CGRP signalling, summarize key clinical evidence for the role of CGRP in migraine headache, including the efficacy of CGRP- targeted treatment, and synthesize what is known about the role of CGRP in the trigeminovascular system. Finally , we consider how the latest findings provide new insight into the central role of the trigeminal ganglion in the pathophysiology of migraine.

1Department of Clinical Sciences, Division of Experimental Vascular Research, Lund University, Lund, Sweden.2Department of Clinical Experimental Research, Glostrup Research Institute, Rigshospitalet, Glostrup, Denmark.3Department of Pharmacology, School of Medicine, University of California at Irvine, Irvine, CA, USA.

*e- mail: [email protected]

https://doi.org/10.1038/ s41582-018-0003-1

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discuss the sites and mechanisms of action of the new CGRP- based migraine therapies. Finally, we consider a new framework within which to think about migraine based on insights from research into the CGRP path-way and consider how this framework could influence future research.

The biology of CGRP transmissionCGRP was first discovered in 1982 (reF.11) and, since then, studies of the trigeminovascular system have told us much of what we know about the role of CGRP in the cranial sensory nerves that are involved in migraine9,12. Multiple components of CGRP transmission are now targeted by migraine therapies (Fig. 2). In the following sections, we review what is known about the various components of CGRP transmission and how they relate to migraine.

CGRP and its releaseCGRP is a peptide neurotransmitter that is produced in peripheral sensory neurons and numerous sites throughout the CNS. CGRP occurs in α and β forms, but the β form has been studied relatively little. The α form is a 37-amino- acid peptide (Fig. 2a) that is synthesized in neurons through tissue- specific splicing of mRNA transcribed from the calcitonin–CGRP gene (CALCA) located on chromosome 11 (reFs11,13). CGRP is generated by cleavage of a pro- peptide precursor and packaged into dense core vesicles for transport to axon terminals and other release sites within the neuron.

In nerves that produce CGRP, the presynaptic termi-nals take the form of focal swellings, called axonal vari-cosities, which occur at regular intervals along the axon to give the appearance of a string of pearls. Upon nerve stimulation, CGRP is released from its storage vesicles via calcium- dependent exocytosis. CGRP release can be stimulated with capsaicin, a component of chili peppers that has consequently been useful as an experimental tool to release and ultimately deplete the peptide from CGRP nerves14,15.

Presynaptic receptors located on trigeminal neu-rons regulate CGRP release. The presynaptic serotonin (5-hydroxytryptamine) receptors 5-HT1B and 5-HT1D inhibit CGRP release16 (Fig. 2c). These receptors are the tar-gets of the triptans (for example, sumatriptan) and medi-ate the effects of these drugs in the relief of migraine4,16–19. A third subtype of presynaptic serotonin receptor, the

5-HT1F receptor, was identified as a target of interest in migraine in 2017 (reF.4). Activation of this receptor also suppresses trigeminal release of CGRP20, and a clinical study has shown that the 5-HT1F agonist lasmiditan has efficacy as a treatment for acute migraine attacks21.

Following its release, CGRP is broken down by metalloproteinases22. Amidation at the carboxyl termi-nus helps to protect the peptide from this breakdown and increase its half- life. This property allows CGRP to spread to receptor targets beyond the release site in a process known as volume transmission23. The pres-ence of CGRP in the blood is generally attributed to spillover from sites of neuronal release, a hypothesis supported by experiments that demonstrated release of trigeminal CGRP into the rat jugular vein24. CGRP has been measured in human blood collected from the external jugular vein, into which extracerebral tissues drain, following trigeminal ganglion stimulation25 (Fig. 3a). Human plasma levels of CGRP are usually in the low picomolar range. The plasma half- life of CGRP in humans following CGRP infusion has been estimated at 7 min during a fast decay phase and 26 min during a slower phase of decay26.

The CGRP receptorThe CGRP receptor is a complex of several proteins, each of which is required for the ligand specificity and func-tion of the receptor (Fig. 2b). Central to the complex is the G protein- coupled receptor calcitonin receptor- like receptor (CALCRL). CALCRL is a member of the secre-tin receptor family (class B G protein- coupled receptor) and is a required element in receptors for CGRP and adrenomedullin (the AM1 and AM2 receptors)27. In order to create a functional membrane receptor with specific affinity for CGRP, CALCRL must form a heterodimer with receptor activity- modifying protein 1 (RAMP1)28. RAMPs are single transmembrane- spanning proteins that alter the pharmacology, functionality and cell traf-ficking of specific G protein- coupled receptors. The ligand- binding domain of the CGRP receptor is located at the interface between RAMP1 and CALCRL29,30. Thus, co- expression of CALCRL and RAMP1 is necessary for a cell to respond to CGRP.

The CGRP receptor complex also includes two cytoplasmic proteins that associate with the CALCRL–RAMP1 heterodimer to mediate signal transduction (Fig. 2b). CALCRL is coupled to a G protein that con-tains the Gαs subunit, which activates adenylyl cyclase and cAMP- dependent signalling pathways13,31. In addi-tion, the CGRP receptor associates with a receptor coupling protein that amplifies G protein activation and is important for optimal signal transduction31.

Receptor- mediated increases in intracellular cAMP activate protein kinase A (PKA), which results in the phosphorylation of multiple downstream tar-gets, including potassium- sensitive ATP channels (KATP channels), extracellular signal- related kinases (ERKs) and transcription factors such as cAMP- responsive element- binding protein (CREB). In cerebrovascular smooth muscle (Fig. 2c), elevation of cAMP upon CGRP activation results in vasorelaxation and dilation of the blood vessel32.

Key points

•Multiplestudieshaveconfirmedthatreleaseofcalcitoningene-relatedpeptide(CGRP)isincreasedduringacutemigraineattacks.

•Inthetrigeminalganglion,CGRPisexpressedinC-fibresanditsreceptorisexpressedinAδ-fibres;thesetypesoffibresareinvolvedindifferentaspectsofpainperception.

•Thetrigeminalganglioniscentraltothetrigeminovascularreflex,whichistriggeredtoprotectagainstvasoconstriction;triggeringofthissysteminpatientswithmigraineleadstotheperceptionofpain.

•Thetrigeminalganglionandduraarenotbehindtheblood–brainbarrier;therefore,theyarelikelytobethetargetsofgepantsandantibodiesinmigrainetreatment.

•CGRPreceptorantagonists,anti-CGRPantibodiesandanti-CGRPreceptorantibodieshaveprovedeffectiveformigrainepainrelief,stronglysupportingthehypothesisthatCGRPhasamajorroleinmigrainepathophysiology.


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An important feature of CGRP signalling is the reg-ulation and desensitization of the receptor following its activation by an agonist33. After CGRP binds to its receptor, CALCRL is rapidly phosphorylated, leading to internalization of the receptor via recruitment of β- arrestins. Transient stimulation by CGRP induces internalization of the receptor to endosomes, from where the receptor can be rapidly recycled back to the plasma membrane. Chronic exposure to CGRP initiates an internalization process that trafficks the receptor to lysosomes for degradation.

Interestingly, there is a second receptor that responds to CGRP, the amylin receptor AMY1, which consists of RAMP1 and the calcitonin receptor27. AMY1 is found in the trigeminal ganglion, although it appears to have a lesser role than the more abundant CGRP receptor34 and it is not blocked by drugs that target the CGRP receptor. At this point, however, whether AMY1 contributes to the mechanisms of migraine and its treatment is unknown.

CGRP receptor antagonistsSeveral small- molecule antagonists have been shown to potently and selectively block CGRP responses in both experimental and clinical studies35,36. Although these molecules are chemically unrelated to one another, they have a shared mode of action, so they are collectively referred to as gepants (Fig. 2c). All gepants that have been tested to date are effective in patients with migraine37–39. Interestingly, a shared feature of the clinically active gep-ants is a high (picomolar) affinity for CGRP receptors of humans and non- human primates relative to their affinity for those of other species. This effect is due to a species- specific residue located at the interface between RAMP1 and CALCRL, indicating that this region is the site of antagonist binding29.

Olcegepant was the first non- peptide CGRP recep-tor antagonist to be discovered40. In cells and in tissue preparations, olcegepant potently blocked the bind-ing of CGRP to its receptor, and in signalling (cAMP production) and functional (vasodilation) assays, it

produced a rightward shift of CGRP concentration–response curves40,41. Olcegepant42 also blocked CGRP responses in vivo in rats43 and was consequently for-mulated for intravenous testing in humans44. Gepants developed subsequently exhibit greater potency than olcegepant and good oral bioavailability; examples include telcagepant45–48, ubrogepant49, MK-3207 (reF.50), BI 44370 TA51 and BMS-927711 (reF.52). However, clin-ical development of several gepants was terminated owing, in part, to a risk of liver toxicity with long- term use6,53 (Fig. 1).

Anti- CGRP and anti- CGRP receptor antibodiesThe first antibodies against CGRP were made shortly after the discovery of the peptide in 1982 (reF.11). These antibodies were initially used for radioimmunoassays, in combination with high- performance liquid chro-matography, to measure CGRP levels in tissues and plasma and for tissue localization of the peptide by use of immunohistochemistry54,55. Specific antibodies are now being used to inhibit the action of CGRP (Fig. 2c): several humanized monoclonal anti- CGRP antibodies (galcanezumab, eptinezumab and fremanezumab) are currently undergoing clinical trials and have proved effective for the prevention of episodic and chronic migraine5,9,10,56,57 (Table 1).

Development of a monoclonal antibody against the CGRP receptor has also been a novel approach to blocking the receptor (Fig. 2c). Erenumab (AMG334) is a human monoclonal antibody that was raised against a fusion protein of the extracellular domains of human CALCRL and RAMP1 that includes the CGRP bind-ing pocket58. This antibody is 5,000-fold more selec-tive for the human CGRP receptor than for other related receptors in the calcitonin receptor family57. Erenumab binds to human CGRP receptors with high affinity and fully antagonizes CGRP responses in vitro and in vivo58. Clinical trials have shown that erenumab is an effective prophylactic therapy for episodic and chronic migraine5,36.

Table 1 | CgrP- related therapies for migraine and other headache disorders

Drug Indicationa Dosing Mechanism Drug development status (September 2017)

Preventive therapy

Erenumab (AMG 334) Migraine prevention in EM and CM

Monthly , subcutaneous Monoclonal antibody against CGRP receptor

Phase III trials complete; registration study published57 and submitted for review to FDA and EMEA

Galcanezumab (LY2951742) Prevention of EM, CM, eCH and cCH

Monthly , subcutaneous Monoclonal antibody against CGRP

Positive results78, now in phase III trials in EM and CM

Fremanezumab (TEV-48125) Prevention of EM, CM, eCH and cCH

Monthly or quarterly , subcutaneous, but intravenous load for cluster headache

Monoclonal antibody against CGRP

Positive results56, now in phase III trials in EM and CM

Eptinezumab (ALD403) Prevention of EM and CM

Quarterly , intravenous Monoclonal antibody against CGRP

Positive results76 in phase III trials in EM; phase III trial in CM ongoing

Acute therapy

Ubrogepant Relief from acute migraine attack

Oral, as needed CGRP receptor antagonist

Positive phase IIb results49; phase III trials ongoing

cCH, chronic cluster headache; CGRP, calcitonin gene- related peptide; CM, chronic migraine; eCH, episodic cluster headache; EM, episodic migraine; EMEA , European Medicines Evaluation Agency. aPrevention is defined as a reduction in headache days.



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Evidence for a role of CGRP in migraineGiven that the trigeminal sensory nerves were already known to have a key role in primary headache disor-ders, the discovery of CGRP in the trigeminovascular system in 1985 immediately suggested that this peptide could be important in migraine pathology59. Moreover, the potent vasodilatory effects of CGRP in cerebral arteries32 fitted with the prevailing view at the time that cerebral vasodilation was an important component of headache.

The presence of CGRP in human trigeminal gan-glia and cerebral arteries was soon confirmed55,60. Interestingly, peptide levels were found to be highest in younger people (aged 20–40 years) and declined with age60, an observation that is consistent with the age- related pattern of headache attacks observed in patients with migraine. Many important milestones in the story of CGRP and migraine have followed (Fig. 1), and we discuss these in the following sections. Together, the findings unequivocally demonstrate that CGRP has a major role in the symptoms of migraine and underscore the effectiveness of blocking CGRP signalling to prevent and abort painful migraine attacks.

Elevated CGRP releaseIn 1990, Goadsby et al. made the seminal observation that CGRP was the only neuropeptide released during the headache phase of migraine attacks; studies of other peptides showed no release in migraine61. In blood sam-ples taken from the external jugular veins of patients, CGRP levels were elevated during the headache (Fig. 3a), whereas levels of other neuropeptides associated with trigeminal and autonomic nerves were unchanged61. CGRP was released during attacks of migraine either with or without aura61. Elevated blood levels of CGRP were also observed during attacks of other primary headache disorders such as cluster headache and chronic paroxysmal headache62,63. In contrast with migraine, however, these two conditions were associated with release of vasoactive intestinal peptide (VIP) in addition to CGRP. Additional studies of patients with migraine have detected elevated levels of CGRP in plasma, saliva and cerebrospinal fluid (CSF) samples63–66. The changes observed in CSF levels might reflect release of CGRP from central projections of the trigeminal neurons, as demonstrated in rats24.

The association of CGRP release with migraine has led to interest in using CGRP as a diagnostic bio-marker66; however, the instability and short half- life of the peptide complicates reliable measurement67. For example, in contrast to the clear results obtained from blood sampling from the external jugular vein during migraine (Fig. 3a), no change was detected in CGRP levels in the peripheral circulation in samples taken at the same time61.

Additional evidence that CGRP has a role in head-ache comes from several clinical studies in which exogenous CGRP was infused intravenously into patients who were prone to migraine attacks68,69. This stimulus induced a lasting, migraine- like headache, suggesting that CGRP has a causal role in migraine symptoms.

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Clinical trials begin to evaluate use of anti-CGRP antibodies for prophylaxis of frequent and chronic migraine

Anti-CGRP antibody (eptinezumab; ALD403), given intravenously, shown to be effective for prevention of episodic migraine (phase II trial)

Ubrogepant, an orally active gepant, shown to be effective in acute migraine without serious adverse effects (phase IIb trial)

Phase III trials of the antibodies eptinezumab, erenumab, fremanezumab and galcanezumab produce positive results for migraine prevention

CGRP antibodies made to measure and localize CGRP in the trigeminal–cerebrovascular system, where it was found to be a potent vasodilator

Discovery of the trigeminovascular reflex: a physiological role for CGRP

Characterization of the multicomponent CGRP receptor that consists of CALCRL, RAMP1 and RCP

Antibodies against CGRP shown to block CGRP responses in vitro and in vivo

First measurement of CGRP released by trigeminal stimulation in humans

Sumatriptan shown to normalize CGRP levels during acute migraine attack in parallel with relief of headache symptoms

Infusion of CGRP shown to trigger migraine attack in patients prone to migraine

Merck files patent for use of CGRP antibodies for migraine treatment

Merck halts development of telcagepant owing to liver toxicity: elevated liver enzymes

migraine prophylaxis

Anti-CGRP antibody (galcanezumab; LY2951742) shown to be effective in episodic migraine without serious adverse effects (phase II trial)

Anti-CGRP antibody (fremanezumab; TEV-48125) shown to be effective in chronic migraine (phase IIb trial)

Antibody targeted to the CGRP receptor (erenumab; AMG 334) shown to be effective in episodic migraine (phase IIb trial)

Expected review of antibody migraine therapies by FDA and European Medicines Agency

CGRP first proposed to play a role in migraine

Discovery of CGRP

Presence of CGRP confirmed in human cerebral vasculature; high levels in children, decreased levels with age

First demonstration in patients that CGRP is released during an acute migraine attack

Demonstration that CGRP release by trigeminal activation is inhibited by triptans

First characterization of compounds that block the CGRP receptor: the gepants

CGRP receptor blocker intravenous olcegepant shown to alleviate headache during a migraine attack

Clinical trials begin to test telcagepant and other gepants in acute migraine

Overview of all clinical trials of gepants to date indicates efficacy in acute migraine with no cardiovascular or other serious adverse effects

after 3-month treatment for

Fig. 1 | Timeline of the key events in the development of drugs that target CgrP for migraine therapy. CALCRL , calcitonin receptor- like receptor ; CGRP, calcitonin gene- related peptide; RAMP1, receptor activity- modifying protein 1; RCP, receptor coupling protein.



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Triptan inhibition of CGRP releaseTriptans have been the mainstay of acute migraine therapy since sumatriptan was first shown to relieve migraine symptoms in the early 1990s4. These drugs are partial agonists of 5-HT1B and 5-HT1D receptors and were originally thought to relieve migraine through vaso-constriction and/or regulation of trigeminal transmit-ter release. In 1993–1994, tests of whether sumatriptan would affect extracranial plasma levels of CGRP in patients during the headache phase of migraine or clus-ter headache16,17,19,62 showed that sumatriptan prevented the increase in plasma CGRP at the same time that it aborted the headache attack (Fig. 3a). This striking find-ing validated the key therapeutic mechanism for triptan drugs and underscored the role of CGRP in migraine.

Efficacy of gepants in migraineAll gepants that have been tested in clinical trials signifi-cantly reduce headache pain and other symptoms asso-ciated with an acute migraine attack36,53 (Fig. 3b). This consistent clinical outcome validates the therapeutic strategy of blocking CGRP receptors to relieve migraine.

The first proof- of-concept study of this approach was a small, double- blind, randomized trial published in 2004 that showed that intravenous olcegepant significantly alleviated symptoms during a migraine attack44. Of the patients with moderate to severe headache, 66% had no or mild headache at 2 h after receiving the dose, thereby meeting the primary efficacy end point.

The encouraging results with olcegepant prompted development of several orally active gepants that were selected for clinical trials. Telcagepant, MK-3207, BI 44370 TA and rimegepant all significantly reduced headache pain after 2 h with similar efficacy47,48,51–54, although the effects of intravenous olcegepant stand out (Fig. 3b). In addition, the gepants significantly reduced photophobia, phonophobia and functional disability at 2 h and provided sustained freedom from pain after 24 h. Testing of gepants proceeded into phase II and phase III clinical trials that demonstrated efficacy in both acute and preventive treatment of migraine10 (Fig. 1). The observed efficacy of the gepants was comparable to that of the triptans but was accompanied by no cardio-vascular or haemodynamic symptoms, which can occur with triptans and make them unsafe for patients with cardiovascular pathologies4. CGRP antagonists do not cause vasoconstriction36, making them safe for use in these patients.

Unfortunately, the momentum in the development of gepants for clinical use in migraine was lost owing to liver toxicity with long- term use10. The first suggestion of this issue arose during a 2009 trial of oral telcagepant for migraine prophylaxis, in which levels of liver enzymes were elevated in some patients who had been taking the medication twice daily for 12 weeks10. The trial was stopped, and within a few years, clinical development of the initial gepants had been discontinued6,53.

Nevertheless, the effectiveness of CGRP antagonists in aborting migraine attacks has led to renewed interest in developing gepants that do not cause liver toxicity. Ubrogepant (MK-1602) and atogepant (MK-8031) are newly developed gepants that are chemically different from telcagepant and the other gepants. These two compounds are currently undergoing clinical testing10. Ubrogepant has completed phase IIb testing and is being evaluated in phase III trials for acute relief of migraine70–72. To date, results indicate that ubrogepant has good efficacy, comparable to that of triptans, with no incidence of elevated liver enzymes or other serious adverse effects10,49. If the newer gepants prove to be safe as well as effective, they will provide a useful alterna-tive to triptans, particularly for patients with migraine and cardiovascular risk factors, patients who do not respond to triptans and patients with triptan- induced medication- overuse headaches6.

Prophylactic efficacy of antibodiesAn alternative to small molecules for blocking CGRP transmission in patients with migraine is the use of selective monoclonal antibodies that bind to either CGRP or the CGRP receptor. The therapeutic goal of this approach is migraine prophylaxis, meaning a reduction in the number of migraine days experienced by patients who have frequent attacks. Four antibodies

Ala

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Cys

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ValVal

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or 5-HT1D

Anti-CGRPantibody

Anti-CGRPreceptorantibody

CGRPreceptor

Gepants

Trigeminal nerve

Protein kinase A

Vasodilation

Fig. 2 | Components of CgrP transmission and sites of action for CgrP- related migraine therapies. a | Amino acid structure of human α- type calcitonin gene- related peptide (CGRP). b | The CGRP receptor complex, which consists of the two integral membrane proteins calcitonin receptor- like receptor (CALCRL) and receptor activity- modifying protein 1 (RAMP1) and two cytoplasmic proteins, receptor coupling protein (RCP) and the α- subunit of the GS protein (GαS). c | The targets for CGRP- related migraine therapies illustrated in a CGRP- containing trigeminal nerve varicosity that innervates a cerebrovascular smooth muscle cell. 5-HT, 5-hydroxytryptamine receptor.



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have been developed and tested to date, including one antibody against the CGRP receptor and three against CGRP itself (Fig. 3c). The anti- CGRP receptor antibody is erenumab (AMG334), a human immunoglobulin G2 (IgG2) monoclonal antibody targeted to the CGRP bind-ing site. The antibody is administered once per month by subcutaneous injection73–75. The three monoclonal anti-bodies targeted to sites on CGRP itself are eptinezumab

(ALD403), a genetically engineered humanized IgG1 monoclonal antibody that is formulated for intrave-nous administration and intended for dosing once every 3 months76; fremanezumab (TEV-48125), a fully humanized IgG2a monoclonal antibody that is admin-istered once per month via subcutaneous injection77; and galcanezumab (LY2951742), a fully humanized IgG4 monoclonal antibody that is also administered subcutaneously on a monthly basis78.

Use of these ant ibodies has successful ly decreased the number of migraine days in episodic migraine10,44–46,49–52,56,57,75–81. Overall, the anti- CGRP antibodies seem to have similar efficacy and safety profiles10. Galcanezumab has also been shown to be effective in treating cluster headache, another disor-der in which CGRP is released during the headache phase62. Phase III clinical trials have been completed for all four antibodies, and the findings are being sub-mitted for review by regulatory agencies for marketing approval (Table 1). The anti- CGRP receptor antibody erenumab is currently under evaluation by the FDA and the European Medicines Agency; if successful, the drug is expected to be marketed in 2018.

Anti- CGRP and anti- CGRP receptor antibodies are particularly suited to prophylactic treatment for migraine, as they have several advantages over other treatment options. Patient adherence and tolerability are both better with the antibodies. The antibodies have a prolonged serum half- life (20–50 days), which means that patients can take the medication infrequently. In addition, antibodies bind to their target site with high affinity and selectivity, thus reducing the potential for unwanted, off- target effects. Furthermore, in contrast to small exogenous molecules such as the gepants, anti-bodies are not processed by the liver, thus avoiding the potential for liver toxicity and hepatic drug interactions. No adverse cardiovascular or cerebrovascular effects of the antibodies under development have been reported to date9,56,57,80,81.

A primary disadvantage of antibodies is that they are not orally active and must be administered by injection. Injection- site reactions, including pain, are common6,56,81, but these reactions are usually mild and transient and are less likely with intravenous administration than with subcutaneous administration. Notably, injection- site reactions, as well as other adverse effects reported during the trials, did not differ between patients who received the antibody and those who received placebo6,80. Overall, clinical trials have shown that the antimigraine antibodies are effective, well tolerated and safe.

Risks of long- term CGRP blockadeDespite the promising data from clinical trials of anti- CGRP and anti- CGRP receptor antibodies, the risks of long- term blockade of CGRP signalling are not known82. Much remains to be learned about the physio-logical and pathophysiological roles of CGRP, but this peptide has effects throughout the body13, and circu-lating antibodies could affect all peripherally accessible sites where CGRP acts.

One concern relates to the role that CGRP plays as a potent vasodilator throughout the vascular system13,83.

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Fig. 3 | Clinical data that demonstrate that CgrP has an important role in migraine headache and its treatment. a | Calcitonin gene- related peptide (CGRP) release during migraine headache. Blood samples were taken from the external jugular veins of patients with migraine during a headache attack and during a headache- free period (control), and blood levels of CGRP were measured61. Successful treatment of headache with sumatriptan was accompanied by lowered blood levels of CGRP. b | Efficacy of the gepants (CGRP receptor antagonists) in treating acute migraine headache. Across several studies, patients with moderate to severe migraine headache received intravenous (i.v.) injection of olcegepant, an orally administered gepant or a placebo. The graph shows the mean percentage of patients who were pain- free at 2 h after drug administration: for olcegepant, 44%44; telcagepant, 27%45,46; MK-3207, 36%50; BI 44370 TA, 27%51; rimegepant, 33%52 and ubrogepant, 25%49. c | Efficacy of CGRP- related monoclonal antibodies in reducing the frequency of attacks in patients with episodic migraine. The graph shows the percentage of patients who had a reduction of >50% in the number of migraine days per month after treatment with either the active drug or a placebo. Data are summarized from published results of phase II clinical trials of the anti- CGRP receptor antibody erenumab (46%)75 and anti- CGRP antibodies eptinezumab (61%)76, galcanezumab (63%)78 and fremanezumab (53%)81 in episodic migraine75–78,143.



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Consequently, chronic reduction of CGRP has the potential to cause cardiovascular pathophysiology, such as hypertension, cardiac dysfunction and epi-sodes of coronary or cerebral ischaemia82,84,85. To date, no cardiovascular adverse effects of anti- CGRP and anti- CGRP receptor antibodies have been reported in up to 6 months of phase III clinical testing6,56,57,80. No patients have developed hypertension related to the treatment. In a study published in 2017, erenumab had no effect on patients with angina pectoris (males aged ~65 years) who were monitored with electrocar-diography while exercising on a treadmill until they reported chest pain86. Of note, the antibody trials have so far mainly included people of middle age who are not at high risk of stroke or acute myocardial infarc-tion. Elderly patients, in whom ischaemic events are of greater concern, are less likely to experience migraine and are therefore unlikely to need the antibody med-ications. Nevertheless, as with any new class of drug, continued monitoring of various patient popula-tions for risks associated with long- term treatment of CGRP- related antibodies is important.

CGRP in the trigeminovascular systemTo better understand how CGRP- targeted therapies affect migraine, we need to define the role of CGRP in the trigeminovascular system, a key component of the pain pathway involved in headache9 (Fig. 4a). As discussed below, good progress has been made in elu-cidating the trigeminovascular locations of CGRP and its receptors and the major functions of this neuropep-tide. Much remains to be learned, but these data, along with recent clinical studies, suggest that long- standing hypotheses regarding the mechanisms of migraine headache need to be re- evaluated. We provide an updated view of migraine pathology and sites of action for anti- CGRP therapeutic drugs.

Expression in the trigeminal ganglionThe presence of a sizeable population of CGRP neu-rons within the trigeminal ganglion signifies a major role for CGRP in trigeminal transmission (Fig. 4b). Immunohistochemical staining with anti- CGRP anti-bodies87–89 and in situ hybridization to localize CGRP mRNA have shown that approximately half of all neu-rons in the trigeminal ganglion express CGRP90. CGRP- positive neurons predominantly have cell bodies with a small to medium diameter, which indicates that they are cells of C- type sensory pain fibres.

Trigeminal CGRP neurons are heterogeneous in that certain subpopulations colocalize with neurons that produce other transmitters, such as pituitary adenylate cyclase- activating polypeptide or substance P54,91, and ion channels, such as the transient receptor potential cation channel subfamily V member 1 (TRPV1) cat-ion channel92. The full extent and implications of these colocalizations have yet to be determined. The majority of CGRP neurons in the human trigeminal ganglion also express 5-HT1D (90% of CGRP neurons) and 5-HT1B (65%) receptors, indicating that the same cells are relevant targets for triptan drugs93. Interestingly, colocalization of CGRP with the CGRP receptor is

rarely observed87, indicating a lack of autoreceptors on CGRP neurons.

Within the trigeminal ganglion, CGRP is also found in thin nerve fibres proximal to the cell bodies, where it colocalizes with the synaptic vesicle marker SNAP25 (reFs87,94). The beaded structure of these fibres suggests that they are CGRP release sites on local branches of CGRP axons (Fig. 4b).

The CGRP receptor can be found in approximately one- third of the ganglion neurons, but these cells are largely distinct from those that contain CGRP (Fig. 4b). Receptor- expressing cells have been identified by use of immunoreactivity to colocalize CALCRL and RAMP1 (reFs87,89) and by visualization of an antibody against the ligand- binding site that spans the CALCRL and RAMP1 subunits88. CGRP receptors are found in the larger neu-rons and thicker fibres, which correspond to Aδ- sensory neurons. This observation corresponds with the findings of a functional study in rats published in 2017, which showed that the CGRP antibody fremanezumab blocks activation of Aδ- type trigeminal neurons but not of C- type neurons95.

Interestingly, certain glial cells within the trigeminal ganglion also express CGRP receptors. In particular, the satellite glial cells that surround neuronal cell bodies can be labelled with either an antibody against the CGRP receptor binding site88 or antibodies against CALCRL and RAMP1, which colocalize on these cells87,89,96. Satellite glia that contain CGRP receptors are often organized around a CGRP- expressing neuron (Fig. 4b), which suggests that neuron–glia communication occurs. The relevance of this type of signalling is not well under-stood, but CGRP has been shown to activate the release of inflammatory cytokines and nitric oxide from gan-glionic glial cells96–100. These substances can in turn enhance CGRP release, thus creating a positive feedback loop within the ganglion101.

Overall, the expression pattern of CGRP and CGRP receptors in the trigeminal ganglion (Fig. 4b) is consist-ent with a model of CGRP signalling in which C- type sensory neurons release CGRP from the soma or local axon varicosities102, and CGRP acts on receptors on Aδ- type sensory neurons and satellite glial cells to modulate pain sensitivity and transmission within the ganglion. These actions of CGRP are likely to be involved in migraine pathophysiology and are targets of anti- CGRP therapies.

Expression in trigeminal nervesBipolar trigeminal sensory neurons release CGRP at peripheral and central nerve terminals27,28. CGRP and CGRP receptors have been visualized in the peripheral and central branches of the trigeminal nerves87. However, the peptide and receptor are localized in distinct fibres, consistent with the expression pattern observed in trigeminal ganglion cell bodies94 (Fig. 4b). CGRP immuno-reactivity is observed in the small- diameter, unmyelinated C- fibres that are known to slowly transmit signals of dull, diffuse pain. By contrast, CGRP receptors are observed in thicker, myelinated fibres indicative of Aδ- type noci-ceptive nerves, which signal sharp, acute pain94. The precise location of receptors in these fibres is not entirely



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clear: receptor immunoreactivity has been observed in axons87,94 and in some of the Schwann cells surrounding the axons (colocalized with immunoreactivity for myelin basic protein)88,89. The close association of C- fibres and Aδ- fibres within the trigeminal nerves suggests that CGRP signalling occurs between the two types of pain fibres, and the presence of CGRP receptors in Schwann cells indicates neuron–glia communication94 (Fig. 4b).

Expression in trigeminal peripheral targetsCerebrovasculature. Shortly after the discovery of CGRP, the peptide was found in nerves that inner-vate cerebral blood vessels and was shown to potently dilate cerebral arteries32,54. Furthermore, the perivas-cular CGRP nerves were shown to be sensory fibres that originated in the trigeminal ganglion54. For >30 years since these discoveries, the cerebrovasculature has provided an important window into trigeminal sensory function and a useful experimental tool for elucidating the mechanisms of CGRP signalling and CGRP receptor pharmacology12.

Trigeminal CGRP nerves that innervate blood ves-sels are located within the vessel wall at the adventitial– medial border and release CGRP from axonal vari-cosities that are separated from the adjacent smooth muscle by a wide cleft (100–500 nm)60,103. CGRP acts on cerebrovascular smooth muscle cells to increase intracellular cAMP, decrease intracellular Ca2+ and cause vasorelaxation, but the peptide has no effect on the cerebrovascular endothelium32,104. CGRP- mediated dilation of cerebral arteries is inhibited by CGRP recep-tor antagonists (gepants) and by antibodies against the CGRP receptor104–107.

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Fig. 4 | CgrP and CgrP receptors in the trigeminovascular system. a | Sites in the trigeminovascular pathway and CNS where calcitonin gene- related peptide (CGRP) and/or CGRP receptors have been identified using immunohistochemistry. The receptor expression pattern resembles that of CGRP, but CGRP does not colocalize with the receptor elements in individual cell bodies or fibres. The only exception is the cerebellar Purkinje cells, which express CGRP and CGRP receptors; however, CGRP is expressed in the cytoplasm, and the receptor elements are expressed in the plasma membrane of cell bodies, axons and dendrites. Red indicates cell bodies that do not express CGRP or CGRP receptors but do express other signalling molecules. b | The relationship between cells and fibres within the trigeminal ganglion that express either CGRP or the CGRP receptor. The question mark indicates a possible interaction. c | CGRP regulation of the trigeminovascular reflex in cerebral arteries on the pial surface of the brain. CGRP mediates the reflex that protects the cerebral circulation from excessive vasoconstriction (left). The importance of CGRP is demonstrated by the effect of vasoconstrictors on cerebral arteries in vivo when the trigeminal innervation is either intact or lesioned (right)108. d | Access to the vessel wall determines the effect of CGRP. The endothelial barrier in the lumen of the vessel prevents CGRP here from accessing the vessel wall (left), so although CGRP is a potent dilator when applied to the abluminal side of isolated arteries, it has no effect when its application is restricted to the vessel lumen (right)43,106.



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The cerebrovascular trigeminal nerves are important for maintaining blood flow to the brain: CGRP increases cerebral blood flow by potently dilating cerebral arteri-oles but not cerebral veins108,109, and this process is criti-cal for protecting the brain circulation by counteracting cerebral artery constriction (Fig. 4c). For example, in subarachnoid haemorrhage, CGRP is released to coun-terbalance blood- induced constriction84. This role was demonstrated by denervating the trigeminal input in cats, which did not alter resting cerebral blood flow but did prolong vasoconstriction of cerebral arteries exposed to a variety of stimuli in vivo (for example, noradrenaline)108. The trigeminovascular vasodilatory reflex in response to vasoconstriction is characteristic of the role that sensory nerves play throughout the body in triggering protective reflexes to harmful stimuli.

One proposed mechanism of antimigraine drugs is inhibition of CGRP- mediated dilation of cerebral arter-ies, which are located on the pial surface of the brain. However, the endothelium in these vessels restricts pas-sage of molecules from the vessel lumen to the outer lay-ers of the vascular wall that contain the smooth muscle and CGRP nerve endings. The hypothesis was tested in isolated rat middle cerebral arteries that were cannulated and luminally perfused, enabling application of drugs to either the endothelium exposed in the lumen or the smooth muscle layers on the abluminal side of the artery. In these experiments, CGRP was effective in relaxing the artery only when applied to the abluminal surface of the vessel43,104,106 (Fig. 4d). Moreover, neither CGRP receptor antagonists (olcegepant and telcagepant) nor anti- CGRP antibodies blocked CGRP- mediated dilation when applied to the lumen; they were effective only when applied to the abluminal side104,106. On this basis, circu-lating CGRP receptor antagonists and antibodies against CGRP and the CGRP receptor, all of which are effective in migraine, do not seem to be able to cross the endothelial barrier to access targets in the cerebrovasculature.

Cranial dura mater. Trigeminal nerves project to the dura mater, where they are involved in meningeal noci-ception and vasodilation110,111. Within the dura, CGRP immunoreactivity is observed in unmyelinated C- fibres, whereas the CGRP receptor components CALCRL and RAMP1 are colocalized in myelinated Aδ- fibres94,112. The role of CGRP receptors in Aδ- fibres is not known, but activation by CGRP could facilitate crosstalk between the two types of sensory nerves to amplify nociceptive signalling or sensitize neurons to such signalling.

CGRP neurons innervate the dural vasculature, including the middle meningeal artery and the superior sagittal sinus94,113,114. As in pial cerebral vessels, CGRP receptor proteins are expressed in the smooth muscle of dural vessels, and CGRP acts here to cause vasodi-lation94,114. However, no endothelial barrier is present in these vessels, allowing CGRP receptor blockers and anti- CGRP antibodies to access the receptors from the lumen.

Unmyelinated fibres of CGRP neurons also termi-nate in nonvascular regions of the dura, where they are activated by noxious and harmful stimuli94,111. The fibres transmit pain messages to the trigeminal ganglion, and

antidromic axon reflexes induce release of CGRP within the dura to increase blood flow to the region. Other neuropeptides are released as well, and together they mediate the process of neurogenic inflammation94,115. Interestingly, CGRP receptors were not found in dural mast cells of humans, although they have been observed in these cells in rats94. Mast cells contain many vasoac-tive and proinflammatory molecules (such as histamine, serotonin, bradykinin and substance P) that are consid-ered important in the neurogenic inflammatory process and have been suggested as having a role in triggering migraine attacks116.

Direct activation of trigeminal afferents in the dura causes a painful headache in humans; this observation was first made in 1940 via stimulation of these vascular nerves during surgery117. Similarities of this headache with migraine headache led to the vascular theory of migraine, which postulates that headaches in this dis-order are triggered by dilation of intracranial blood vessels118. However, this view is now being disputed, particularly in light of studies that have demonstrated that cerebral and extracranial meningeal arteries are not dilated during a migraine attack119 and a headache does not result from their dilation120.

The neurogenic inflammation in the dura has also been proposed as the trigger for migraine attacks116. However, numerous drugs that block the plasma protein extravasation component of neurogenic inflammation in the dura of animals have been tested in clinical trials but have not exhibited antimigraine efficacy121. Moreover, CGRP does not induce neurogenic inflammation in humans or rodents but mediates only the vasodilatory aspect of inflammation121. Thus, the dura does not seem to be the primary site at which CGRP- targeted thera-pies act to relieve migraine headache. Indeed, current thinking discounts dural mechanisms as the trigger of migraine and focuses on neural mechanisms122,123. Nevertheless, no access barrier within the dura or its vessels excludes CGRP- related drugs; therefore, the possibility that the actions of the drugs in these loca-tions contribute to the overall therapeutic effect cannot be ruled out.

Expression in trigeminal central targetsSpinal trigeminal nucleus and spinal cord. CGRP is abundant in the central terminals of primary trigeminal afferents. CGRP- positive axons terminate in the spinal trigeminal nucleus (STN) in the brainstem and in the upper cervical levels of the spinal cord (C1 and C2), nota-bly in laminae I and II of the dorsal horn124–128. At these central sites, trigeminal input is relayed to second- order neurons of the pain pathway that project via the brainstem and midbrain to higher cortical pain regions122,129,130.

As in the peripheral branches, the central CGRP- immunoreactive fibres are thin, unmyelinated axons that contain pearl- like structures that are vesicular- localized CGRP storage sites. CGRP colocalizes with synaptic ves-icle protein, which indicates the presence of CGRP at vesicle release sites124. These fibres primarily derive from the ipsilateral trigeminal ganglion. The CGRP receptor components CALCRL and RAMP1 are found in thicker fibres that express a marker for Aδ- fibres124.



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In the human STN, the highest density of CGRP- immunoreactive fibres is observed in a network around fibre bundles in the superficial laminae124. In the cervical spinal cord, CGRP labelling is most intense in laminae I and II and is weaker in lamina V (reF.124). Interestingly, no neuronal cell bodies in these regions express CGRP or CGRP receptors; these elements are observed only in nerve terminals. This observation suggests that CGRP acts to modify the signalling of Aδ- fibre afferents.

Ascending pain pathways and other CNS sites. The loca-tion of CGRP and CGRP receptors throughout the CNS has been extensively mapped131–133. Both are expressed at numerous sites associated with pain processing and other functions associated with migraine symptoms, such as nausea, photophobia and phonophobia122. However, the blood–brain barrier (BBB) makes these sites unlikely as targets that mediate the therapeutic actions of CGRP- related antibodies. Detailed studies of the BBB, in which the Evans blue albumin permeability method and the quantitative permeability surface area method of Fenstermacher were used134–136, have demonstrated that permeability was >30-fold greater in the trigeminal ganglion than in the CNS and the trigeminal nucleus caudalis91,137. The limited passage of the antibodies (<0.01%) and some gepants (2%) into the brain speaks

against the CNS as a primary site for their therapeutic action9. Pharmacologically, the amount of drug necessary to inhibit CGRP signalling in the CNS is not achievable; therefore, the main target must be outside the BBB.

CGRP and/or CGRP receptors are expressed in several CNS sites that lack a BBB, such as the area pos-trema132, which might have a role in migraine- associated symptoms such as nausea and vomiting. Therefore, these sites could be accessible to CGRP- related drugs. Furthermore, trigeminal nerves have other projections outside the BBB that express CGRP and its recep-tors (Fig. 4a), such as the sphenopalatine ganglion, which facilitates crosstalk between the sensory and parasympathetic systems.

Updated view of migraineFindings regarding CGRP and the effectiveness of CGRP- targeted therapies in migraine offer new insight into the pathophysiology of migraine headache. However, questions remain about how CGRP mecha-nisms fit into the overall theory of migraine. Although there are many causes of headache, migraine is a primary neurological disorder with recurrent attacks that seem to originate in the brain57,122,123,138: several CNS areas are active during the prodrome, aura and headache phases of migraine122,123,138. Nevertheless, several migraine therapies, including certain triptans, gepants and the CGRP- related antibodies, have little or no ability to cross the BBB, so they seem to act outside the CNS106,139. Furthermore, although sumatriptan relieves migraine headache, it does not inhibit ongoing activation of brain-stem areas that evidence suggests are so- called migraine generators123,140.

A peripheral site of action for CGRP- related therapies fits with traditional theories of migraine in which head-ache is thought to be triggered by either vasodilation of cranial arteries or neurogenic inflammation in the dura. However, these theories have not held up against the latest clinical observations119,121,141, indicating that another peripheral site must be involved in generating and maintaining the headache phase of migraine.

The trigeminovascular pathway is known to be acti-vated during a migraine attack, and the brain interprets signalling in this pathway as headache pain, as Wolff dra-matically demonstrated in his surgical patients117. Studies of CGRP also clearly indicate that peripheral aspects of the trigeminal system have an important role during the headache phase. CGRP is released into the cranial circu-lation during an attack, probably from neurons and fibres of the trigeminal ganglion, where 50% of the neurons contain CGRP and there are no barriers to the periph-eral circulation28. Normalization of CGRP levels in sam-ples from the jugular vein corresponds with resolution of the headache upon administration of triptans23,61,142. These observations demonstrate that trigeminal CGRP release is a good indicator of the attack, but the exact role of this process in headache generation is not clear. Given that the trigeminal ganglion is central to the trigemino-vascular pain pathway, speculation that blocking CGRP transmission within the trigeminal ganglion is sufficient to abort or prevent the debilitating symptoms of migraine seems reasonable.

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Fig. 5 | Proposed involvement of the trigeminal ganglion in migraine headache and mode of action of CgrP- targeted therapies. The migraine attack is initiated in the CNS, probably involving regions in the dorsal pons, hypothalamus and thalamus (putative migraine generators)123. The trigeminal ganglion is activated, triggering stimulation of afferents of the trigeminovascular pathway that project to the spinal cord and convey the sensation of head pain to the CNS, initiating the headache phase. These afferents terminate behind the blood–brain barrier in the spinal trigeminal nucleus and the dorsal horn of spinal levels C1 and C2. Second- order neurons then project the pain signal through ascending pain pathways to thalamic and cortical regions that perceive headache pain. We propose that these signals can run in both directions, thereby creating a cycle in which the trigeminal ganglion acts as an amplifier of migraine headache pain owing to persistent activation of calcitonin gene- related peptide (CGRP) circuits within the trigeminal ganglion that reverberate via crosstalk between sensory neurons (C- fibres and Aδ- fibres) and satellite glia (Fig. 3b). Blocking CGRP signalling within the trigeminal ganglion would abort the migraine headache and/or prevent it from occurring. New CGRP- targeted therapies, such as anti- CGRP antibodies, the anti- CGRP receptor antibody and gepants, can all access the barrier- free trigeminal ganglion to exert this action.



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ConclusionsOn the basis of what we have learned about the CGRP system and the effectiveness of new CGRP- related drugs, the painful, lasting headache could be a conse-quence of dysfunctional signalling within the trigem-inal ganglion. To stimulate further discussion within the field, we propose a migraine model (Fig. 5) in which during the headache phase, persistent activation of the trigeminal ganglion and its local CGRP circuits ampli-fies the strength and duration of signalling in central trigeminal afferents that convey the sensation of head pain to the CNS. Thus, the ganglion might act as an amplifier that promotes intense and long- lasting pain.

However, this model does not explain how the ganglion is initially activated during migraine. Nevertheless, on this basis, the most likely targets of therapeutic CGRP antibodies and CGRP receptor blockers are within the trigeminal ganglion itself, where they would act to abort and/or prevent amplification of the pain signals. Much remains to be learned about how CGRP transmission in the trigeminal ganglion contributes to migraine attacks, but the development of effective drugs that act on this target is a major milestone for patients with migraine.

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Author contributionsAll authors researched data for the article, made substantial contributions to discussion of the content and reviewed and/or edited the manuscript before submission. L.E. and D.N.K. wrote the manuscript.

Competing interestsThe authors declare no competing interests.

Publisher’s noteSpringer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Reviewer informationNature Reviews Neurology thanks H.-C. Diener, A. Rapoport, A. Russo and C. M. Villalón for their contribution to the peer review of this work.



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