Background
Cluster headache is one of the most painful disorders in the world, with a prevalence of around 0.1% and a clear male predominance [
1]. It is characterized by excruciating unilateral orbital, supraorbital or temporal pain attacks, with accompanying cranial autonomic symptoms or restlessness/agitation. The attacks last between 15 and 180 min but may occur up to 8 times per day and take weeks to months to remit. In some patients, the cluster headache presents as a chronic form, with attacks occurring for > 1 year without remission, or with remission periods lasting < 3 months [
2]. The clinical features of cluster headache in Asian populations differ from those in Western populations, including a lower prevalence of chronic cluster headache, higher male/female ratio, and lower frequencies of restlessness or aura [
3,
4]. Mechanisms underlying these ethnic differences remain unclear.
Despite the tremendous impact on the sufferers, detailed pathogenesis underlying cluster headache remains enigmatic. Because cluster headache has significant familial aggregation, attempts have been undertaken to determine its underlying genetic architectures. Hypothesis-driven approaches using candidate gene association studies, however, have not identified replicable signals. The first genome-wide association study (GWAS) investigating 99 Italian patients with cluster reported suggestive associations with genetic variants in
ADCYAP1R1 (ADCYAP receptor type I) and
MME (membrane metalloendopeptidase) [
5]; however, these findings were not replicated subsequently. Two recent studies provided the first evidence to demonstrate genome-wide significant variants contributing to the predisposition of cluster headache in European cohorts [
6,
7]. The Dutch and Norwegian study identified rs11579212 near
RP11-815 M8.1, rs6541998 near
MERTK (MER Proto-Oncogene, Tyrosine Kinase), rs10184573 near
AC093590.1, and rs2499799 near
UFL1 (UFM1 specific ligase 1)/FHL5 (four and a half LIM domains 5)[
6] whereas the combined United Kingdom (UK) and Swedish cohorts identified rs113658130 near
LINC01877/SATB2 (SATB homeobox 2), rs4519530 in
MERTK, rs12121134 near
LINC01705/DUSP10 (
Dual Specificity Phosphatase 10), and rs11153082 in
FHL5 to be associated with cluster headache [
7]. Considering the inter-ethnic variability of clinical characteristics [
3], it is uncertain whether these novel loci are replicable in other populations.
To interrogate the genetic architecture of cluster headache in Asians, we performed a two-stage GWAS in a total of 734 clinic-based patients and 9,846 population-based controls. We also established polygenic risk score (PRS) models to differentiate patients from controls and conducted downstream analyses to investigate genes and potential pathogenic mechanisms of cluster headache.
Discussion
We identified three susceptibility loci, in
CAPN2, MERTK, and SATB2, as well as one suggestive locus at
CYP2C18/
CYP2C19 at genome-wide level in patients with cluster headache in Taiwan. To the best of our knowledge, this is the first GWAS of cluster headache performed in Han Chinese and the first in Asians. While replicating the susceptibility genes recently identified in GWASs in patients of European ancestries [
6,
7] suggested the validity of our study, we also identified novel genes implicating potential inter-ethnic differences. The association effect sizes are relatively large and similar to those observed in European GWASs [
6,
7]. These results suggest that some phenotypes of cluster headache might be driven by these selected loci with large effect size, although further studies are needed to explore the genotype–phenotype association. In addition, several other risk loci identified in previous studies are of suggestive GWAS significance in our samples, suggesting that future studies with larger sample sizes might validate the associations of these loci with cluster headache. Moreover, the superior discriminative capacity of genome-wide PRS than the PRS composed of known cluster headache-associated loci suggests that additional loci with smaller effect sizes might also contribute to the genetic basis of cluster headache. Nevertheless, the clinical utility of PRS remains to be explored.
The novel gene
CAPN2 identified in our study encodes calpain 2, a calcium-regulated non-lysosomal thiol-protease involved in cytoskeletal remodeling and signal transduction. Calpain 1/2 have been known to mediate Ca
2+ influx and mediate degradation of suprachiasmatic nucleus (SCN) circadian oscillatory protein (SCOP) [
40] in SCN neurons, which may contribute to coordinated regulation of circadian rhythms. In addition, calpain 1/2 play opposite roles in retinal ganglion cell (RGC) degeneration induced by ischemia/reperfusion injury [
41], while degeneration of RGCs could lead to impaired circadian rhythmicity. Another potential implication of
CAPN2 may be that the most commonly used preventive drug for cluster headache, the L-type calcium channel blocker verapamil, has been known to abrogate calpain activation [
42]. Furthermore, calpain was found to mediate capsaicin-induced ablation of transient receptor potential vanilloid subtype 1 (TRPV1)-positive trigeminal afferent terminals [
43], which may mediate the release of calcitonin gene-related peptide (CGRP) and contribute to the pathogenesis of cluster headache.
We successfully replicated
MERTK and
SATB2 identified in previous cluster headache GWASs [
6,
7].
MERTK encodes a protein belongs to the MER/AXL/TYRO3 receptor kinase family. In addition to regulating microglia-mediated neuroinflammation and astrocyte-mediated neuronal synaptic remodeling proposed in previous studies [
6,
7].
MERTK functions in the retinal pigment epithelium as a regulator of photoreceptor phagocytosis, which is a circadian-regulated process indispensable for vision [
44]. Mutations of
MERTK cause degeneration of photoreceptors, which in turn lead to the loss of photic and circadian control and reduced production of melanopsin mRNA in RGCs [
45]. As melanopsin-expressing RGCs are responsible for circadian photoreception and project to SCN and hypothalamus [
46],
MERTK may thus indirectly participate in the pathogenesis of cluster headache. In addition, enrichment analysis showed significant expression of
MERTK in pituitary gland, which could potentially contribute to the altered hormonal expression in cluster headache [
47].
SATB2 encodes a DNA binding protein that specifically binds nuclear matrix attachment regions and involves in transcription regulation and chromatin remodeling. Previous GWAS suggested that it may be associated with hypothalamic dopaminergic neurons and structures responsible for nociceptive processing [
6,
7].
SATB2 was also known as an important transcription factor of RGCs in primates [
48]. In addition, both GWAS and gene-based association analysis suggested
CYP2C18 as a potential novel susceptibility gene for cluster headache. Interesting, CYP2C18 is a cytochrome P450 monooxygenase involved in retinoid metabolism [
49], while retinoic acid is a molecular trigger of RGC hyperactivity [
50]. Taken together, the top implicated genes in our study may modulate the function of RGCs; however, how this could be involved in cluster headache pathogenesis particularly circadian rhythmicity requires further validation.
Gene-based association testing also identified several possible additional loci, among these,
ANAPC1 has also been identified in another cluster headache GWAS using gene-based analysis, with enriched expression in the brain, particularly neurons [
7]. In addition, among the genes that may be potentially affected by variants in
MERTK, i.e., with significant eQTL expression,
TMEM87B (
transmembrane protein 87B) and
FBLN7 (
Fibulin 7) have been identified in prior GWASs via gene-based analysis or eQTL analysis [
6,
7], suggesting their involvement in cluster headache. Moreover, we found evidence of suggestive association among previously suggested loci to be involved in cluster headache, including
LINC01877, LINC01705, ADCYAP1R1,
and MME. Although these genes are functionally plausible for the pathogenesis of cluster headache, future studies with larger sample sizes are needed to validate these findings.
We found considerable heritability of cluster headache and migraine and significant genetic correlation between these two primary headache disorders, which corroborates with the clinical observations that both disorders exhibit features of trigeminovascular activation and respond to similar treatments such as triptans or CGRP monoclonal antibodies. Some previously reported migraine loci were also found to have suggestive GWAS significance in our current sample, which may contribute to the share biology between migraine and cluster headache. In addition, pathway analyses suggested that biological processes associated with synaptic transmission or immune responses may be involved in the pathogenesis of cluster headache. In fact, immunological processes have long been considered important in the pathogenesis of migraine and we have recently found
HLA class I alleles are associated with clinic-based migraine [
51].
Our study has limitations. First, the sample size is relatively small. However, the successful validation of the findings of previous GWASs and replicable signals in both stages of our GWAS after stringent quality control suggest that our findings are unlikely spurious. Second, the identified variants might not be the true causal variants and it remains unknown whether the expression of the implicated genes is truly altered in patients even though eQTL analysis suggested that these variants could affect gene expression in certain tissues. Finally, although the implicated genes were plausibly relevant to the pathogenesis of cluster headache in in silico functional analysis, we were unable to validate the function of these genes in vivo at the current stage owing to the limitation of the availability of biological samples such as brain tissues or retina from the patients. Further in-depth functional analysis at molecular level, at least in a subset of the patients, are needed the increase the credibility of the findings..
Publisher’s Note
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.