Background
Migraine is one of the most common debilitating disorders, affecting 324 million people worldwide [
1]. Even though the acute treatment of migraine has greatly improved with the development of 5-HT
1B/1D receptor agonists (triptans), a substantial percentage of patients do not benefit from oral triptan formulations [
2,
3]. Triptans are contraindicated in patients with cerebrovascular disease, cardiovascular disease, poorly controlled hypertension, severe hepatic or renal impairment and certain forms of migraine; triptans may also induce serotonin syndrome when taken in combination with selective serotonin-reuptake inhibitors [
4]. In the majority of patients, the migraine attack is accompanied by cutaneous allodynia [
5] and this phenomenon has been reported to be associated with a lack of efficacy of triptans [
6]. Even though these findings were not supported by later studies [
7‐
9], they emphasize the importance of developing alternative treatment options for patients that do not benefit from triptans.
The neurotransmitter calcitonin gene-related peptide (CGRP) plays a crucial role in the pathophysiology of migraine. Infusion of CGRP causes migraine-like disorders or even migraine without aura [
10], and several CGRP receptor antagonists have been shown to be effective in the acute treatment of migraine [
11‐
13]. It is well known that activation of the trigeminal nerve system induces a release of CGRP [
14] and our group has recently demonstrated that this release could be almost completely abolished by destroying primary trigeminal afferents with neonatal capsaicin treatment [
15].
Capsaicin activates the heat and pH-sensitive ion channel Transient Receptor Potential Vanilloid 1 (TRPV1), which seems to be involved in the pathophysiology of migraine [
15‐
18]. TRPV1 is expressed on trigeminal nociceptors [
19,
20], which innervate the dura mater and the meningeal vascular system [
21]. Activation of TRPV1 causes release of CGRP from trigeminal nerve terminals [
22‐
24] and neurogenic inflammation within the meninges [
25], possibly initiating migraine attacks. Accordingly, the anti-migraine drug sumatriptan was recently shown to block trigeminal TRPV1 channels [
26].
However, the effectiveness of specific antagonism of the TRPV1 channel in the treatment of migraine remains unresolved. Even though one antagonist, SB-705498, was shown to suppress and reverse sensitization upon dural inflammation [
27], a clinical trial testing the compound in migraine patients has been terminated early due to lack of efficacy [
28]. Another TRPV1 antagonist, A-993610, was recently shown to be ineffective in different animal models of migraine [
29]. Here, we investigate the effectiveness of two TRPV1 receptor antagonists in blocking trigeminal activation, as measured by expression of the immediate early gene
c-fos [
30] and in preventing CGRP release, both common
in vivo models of migraine.
Discussion
In this study we applied two
in vivo models of migraine to test the efficacy of two TRPV1 antagonists. It is commonly believed that activation of the trigeminal nerve system and subsequent release of CGRP from trigeminal fibres, which leads to vasodilation within the meninges, plays a crucial role in the development of migraine attacks. This is supported by the fact that several CGRP receptor antagonists are effective in the acute treatment of migraine [
11‐
13]. We have shown here that antagonists of the TRPV1 ion channel could be effective in blocking this trigeminal activation that leads to the release of CGRP.
First, we tested the effect of two TRPV1 antagonists on the stimulus-induced up-regulation of the immediate early gene
c-fos in the trigeminal brain stem complex. We selected the so-called inflammatory soup (IS) as activating stimulus, which is a well-established model to study the mechanisms underlying the sensitization of trigeminal primary afferent neurons. This central sensitization has been considered to be the cause of certain headaches and is almost certainly the underlying mechanism for allodynia, which often accompanies migraine attacks [
36,
37]. Our group has shown that IS activates the trigeminal nerve system when administered intracisternally as demonstrated by CGRP release [
32]. Here, we confirm these findings by showing that intracisternal administration of IS leads to a pronounced up-regulation of
c-fos LI within the TNC. The amount of
c-fos expression in response to IS was found to be considerably less than what has previously been reported as the response to capsaicin [
31]. As IS predominantly acts through the sensitization rather than direct activation of nociceptive ion channels, this result was not unexpected. Elevated
c-fos in the TNC after stimulation of the dura with IS has been reported previously and the results correspond well with the findings of this study [
38,
39]. However, Edelmayer et al. [
39] have shown sumatriptan to be effective in blocking
c-fos up-regulation, which does not correspond to our results. This discrepancy is likely due to lower concentration of inflammatory mediators and higher concentration of sumatriptan used by Edelmayer et al. We have shown in this and earlier reports that the lower sumatriptan dose administered here is already effective as it blocks both capsaicin and IS-induced CGRP release [
32]. We do not propose a direct effect of sumatriptan on TRPV1 channels but rather an inhibition of CGRP release by the drug. However, sumatriptan does not reverse sensitization of already sensitized central trigeminal neurons [
40], thus explaining the absence of an effect on the IS-induced
c-fos up-regulation in this study. The TRPV1 antagonist JNJ-38893777 in its highest dose (3 mg/kg) decreased the IS-induced
c-fos LI significantly, in some areas even to the level of control animals not treated with IS. The second antagonist JNJ-17203212 showed remarkable efficacy in all levels of the TNC at 30 mg/kg, completely abolishing
c-fos up-regulation. To corroborate the above results, we tested the two TRPV1 antagonists for their ability to block stimulus-induced CGRP release into jugular vein blood. Capsaicin was used as the stimulus as this substance has been used in many preclinical migraine drug studies previously [
41,
42]. In addition, the use of capsaicin allowed us to demonstrate the efficacy of the two antagonists in blocking the TRPV1 channel. Injection of capsaicin into the carotid artery caused a significant increase in jugular CGRP levels that was sustained for 15 min. The TRPV1 antagonist JNJ-38893777 at 3 mg/kg was more effective than sumatriptan and returned CGRP levels close to baseline. The TRPV1 antagonist JNJ-17203212 was effective in reducing capsaicin-induced CGRP release at all three concentrations. These results demonstrate the specificity of the two compounds against the TRPV1 channel and suggest that the IS-induced up-regulation of
c-fos was also due to a block of TRPV1.
The specificity of the two antagonists used in this study has been tested extensively
in vitro and the results are summarized in Table
1. Both compounds were tested against a number of TRP channels as well as against a broad panel of other non-related receptors and channels. JNJ-17203212 displayed some weak inhibition of TRPM8 that remained far below the activity at the TRPV1 channel, but was not active at any of the other receptors and channels [
43]. JNJ-38893777 was not active at any of the tested TRP channels but displayed some weak activity at the human Cholecystokinin 1 receptor (p
K
i 6.1), the human Adenosine 3 receptor (p
K
i 6.2) and a rat cerebral cortex sodium channel (p
K
i 6.1). However, these activities were far lower compared to the inhibition of hTRPV1 (p
K
i 8.0). In conclusion, while we cannot exclude the possibility that the effect of the two antagonists on
c-fos expression could also be mediated by additional targets, a significant effect seems unlikely given the higher specificity of the compounds against the TRPV1 channel and the TRPV1-specific block observed in the CGRP experiments.
Table 1
Pharmacological selectivity of JNJ compounds
pIC50 (capsaicin) | 8.08 | 7.19 |
pIC50 (low pH) | 8.13 | 7.8 |
pK
i (hTRPV1) | 8.0 | 7.3 |
Activity against related TRP channelsa
| no activity (pIC50 < 5) | weak activity against cTRPM8 (pIC50 < 6) |
Activity against non-related channels or receptors | weak activity against: | no activity |
human Cholecystokinin 1 receptor (pK
i 6.1) |
human Adenosine 3 receptor (pK
i 6.2) |
rat cerebral cortex sodium channel (pK
i 6.1) |
TRPV1 is well known for its role in nociception and sensitization and has been widely studied in the peripheral nociceptive system [
44]. In the central nervous system, TRPV1 expression has been shown in the TNC and in numerous other areas [
26,
45‐
47], many of which are involved in the processing during headaches [
48]. TRPV1 is expressed by a majority of CGRP-releasing trigeminal fibres that innervate the meningeal vascular system [
15,
49]. Stimulation of these trigeminal afferents through activation of TRPV1 causes a CGRP-mediated increase in dural blood flow [
22,
24,
49] and activation of second order neurons in the TNC [
33], and these mechanisms could be involved in the initiation of migraine attacks. Because activation of the trigeminal nucleus is essential during the pain phase of a migraine attack, we decided to study the effect of TRPV1 antagonism in this anatomical region. While we cannot exclude the possibility that the TRPV1 antagonists used in this study also acted on sites other than the TNC, our
c-fos data point to an efficacy of the two compounds in that particular area. This is in good agreement with previous data, showing that functional antagonism at the TRPV1 receptor may modulate neurotransmission in the TNC [
50].
Research on the importance of the TRPV1 channel in the pathophysiology of migraine is, however, inconclusive. A growing body of evidence seems to support a role of TRP channels in general and TRPV1 in particular in the pathomechanisms of headaches [
16,
51]. A recent study investigating single nucleotide polymorphisms among the Spanish population identified the TRPV1 and TRPV3 genes as likely candidates for contributing to an increased genetic susceptibility to migraine [
17]. The well-known migraine trigger ethanol has been shown to induce neurogenic vasodilation via a TRPV1-mediated release of CGRP [
24]. Furthermore, TRPV1 channels in the TNC or in dissociated trigeminal neurons were shown to be inhibited by the common migraine drug sumatriptan [
26]. This is in line with the findings in this study that show reduced trigeminal activation upon block of TRPV1 channels. Confirming these observations, electrophysiological
in vivo studies have shown that a direct inhibition of the TRPV1 channel using the TRPV1 antagonist SB-705498, as well as the functional antagonism using the TRPV1 agonist olvanil, which induces a long-lasting neuronal desensitization, suppress neuronal activity in the trigeminocervical complex following nociceptive dural stimulation [
27,
50]. A recent study showed that CGRP increases TRPV1 expression in the trigeminal nociceptive system [
18]. This is strengthened by the observation that CGRP levels in jugular vein blood of human patients are elevated during migraine attacks [
52]. Taken together, these findings emphasize the importance of the interplay of TRPV1 and CGRP in migraine-related processes. However, the interpretation of these findings is still controversial as conflicting data exists. In this context, electrophysiological studies using another TRPV1 antagonist, A-993610, failed to show alleviating effects in different animal models of migraine [
29]. The authors measured trigeminal firing, induced by electrical stimulation of the middle meningeal artery, neurogenic dural vasodilation and mechanically induced cortical spreading depression. The failure of A-993610 to be effective is in contrast to the results of this study. We used two different models of trigeminal activation and two completely different TRPV1 antagonists, both of which showed significant effects. The published IC
50 value for A-993610 [
53] seems to lie in a similar range compared to the two compounds used here. However, no further information on pharmacokinetics or bioavailability for this particular compound could be found. In light of the promising data reported here, it seems that it might be warranted to test the two compounds described here in an experimental paradigm similar to that described by Summ et al. [
29] as well as in further clinical studies.
A major problem with the clinical use of early developed TRPV1 antagonists was a significant hyperthermia, which in several human patients lasted for several days and could exceed 40 °C [
54,
55]. However, more recently developed antagonists have been shown to avoid this severe side effect [
56]. Nevertheless, the use of TRPV1 antagonists for the treatment of acute headache or migraine is controversial. The promising TRPV1 antagonist SB-705498 was shown to be effective in preventing and reversing sensitization of responses to electrical stimulation, induced by topical application of IS onto the dura mater of cats [
27]. Conflicting results were, however, obtained in human patients. A clinical trial using this compound to treat acute migraine has been terminated early due to a lack of efficacy [
28]. SB-705498 had previously failed in reducing capsaicin-evoked hyperalgesia and had only minimal effects on capsaicin-induced flare [
57], and it is not clear if the properties of this compound are representative of all TRPV1 antagonists. Direct comparison revealed an overall higher TRPV1 affinity of the compounds used in this study compared to SB-705498. Both JNJ compounds have higher p
K
i and in some paradigms higher pIC
50 values [
43]. JNJ-17203212 also displays a longer half-life of 7.4 h [
58] compared to 3.1 h for SB-705498 [
59]. Furthermore, oral bioavailability exceeds that of SB-705498 by 14 %. JNJ-38893777 was recently shown in a human study to be well tolerated without causing serious adverse events and to be suitable for further clinical development [
60]. Taken together, these findings might suggest that the compound SB-705498 and the dosage used in the previous clinical trial were not optimal. Additional clinical studies with other TRPV1 antagonists should therefore be conducted before definitive conclusions on the role of TRPV1 in migraine can be drawn.
Competing interests
The study was funded by a research grant to UR from Janssen Research & Development, L.L.C.
UR has received honoraria for the participation in advisory boards, oral presentations or contributions to clinical trials from Pharm Allergan, Amgen, Almirall, Autonomic technologies, Astra Zeneca, Berlin Chemie, Böhringer Ingelheim, Co-Lucid, ElectroCore, Haas & Health, MSD Sharp and Dohme, Janssen Cilag, GSK, Pfizer. UR has no ownership interest and does not own stock in any pharmaceutical company.
Authors’ contributions
JEM carried out c-fos experiments, participated in the data analysis and drafted the manuscript. JH carried out c-fos experiments, participated in the data analysis and reviewed the manuscript. SRC and AW provided the antagonists, participated in the design of the study and reviewed the manuscript. LN was involved in the design of the study, carried out the CGRP assays and contributed to the data analysis. SSH carried out the CGRP experiments and contributed to the data analysis. UR conceived, designed and coordinated the study, participated in the data analysis and reviewed the manuscript. All authors read and approved the final manuscript.