Clinical trial NCT06549270: Biochemical Profiling of Migraine Patients
Official title: Unraveling the Spectrum of Migraine Resistant to Treatments: Searching for Novel Biological PHEnotypes and theRApeutic Approaches (SPHERA Project)
Study type: OBSERVATIONAL
Overall status: RECRUITING
Conditions: Migraine Disorders
Chronic Migraine
Episodic Migraine
Keywords: migraine
anti CGRP mABs
calcitonin gene related peptide
endocannabinoid system
Lead sponsor: IRCCS National Neurological Institute "C. Mondino" Foundation
Sponsor class: OTHER
Collaborators: name: University of L'Aquila
First posted: 2024-08-12
Enrollment: 100
Sex: ALL
Minimum age (years): 18
Maximum age (years): 75
Standard ages: ADULT
OLDER_ADULT
Observational model: CASE_CONTROL
Time perspective: PROSPECTIVE
Sampling method: PROBABILITY_SAMPLE
Study population: Subjects with chronic or high frequency episodic migraine attending the outpatient clinic of the Headache Science \& Neurorehabilitation Unit of the IRCCS Mondino Foundation (Pavia, Italy) and the Neurology Department of the University of L'Aquila (Avezzano, Italy).

Brief summary: Aim of the study was to assess a potential dysfunction of the endocannabidiome system (eCBome) in migraine patients. Migraine patients who will undergo preventive therapy with monoclonal antibodies directed against the calcitonin gene related peptide (mAbs) will be evaluated through a deep phenotyping of peripheral neurochemical biomarkers (eCBome, neuropeptides, cytokines and kynurenine levels, and microRNAs expression).

Primary aim is to assess baseline differences among those patients who achieved a reduction of monthly migraine days \>/= 50% after three months of tretament (namely Responders) and those who did not (namely Non-responders).

Detailed description: Previous evidence showed that endocannabidiome system (eCBome) is altered in migraine patients demonstrating: i) altered gene expression of catabolizing enzymes (MAGL and FAAH) in patients with episodic and chronic migraine compared to healthy controls; ii) altered peripheral levels of the endocannabinoid-like lipid palmitoylethanolamide (PEA) with evidence of increased PEA levels during the acute migraine phase.

Despite the high effectiveness and tolerability of mAbs monoclonal antibodies directed against the Calcitonin gene related peptide (mAbs), evidence from RCTs and real-life studies demonstrates that mAbs fail in 40% of patients. These patients may bear a non CGRP- dependent phenotype, potentially linked to eCBome dysfunction.

Primary aim is to perform a deep phenotyping of the whole cohort of migraine patients comparing the subgroups of those patients who will be Responders to mAbs treatment (namely those patients who achieved a reduction of monthly migraine days \>/= 50%) compared to the Non-Respoder group (namely those patients who achieved a reduction of monthly migraine days \< 50%) .

Neuropeptides, microRNAs, inflammatory cytokines, and kynurenine metabolites will be evaluated. These findings will allow the identification of a multibiomarkers panel signature of migraine patients resisting to specifically targeted preventive treatments and potentially unveiling other molecular targets.

STUDY DESIGN:

This study is part of the SPHERA project with funding from the Italian Ministry of Health (GR-2021-12372429). Patients will be enrolled from those attending the outpatient clinic of IRCCS Mondino Institute (Pavia) and Neurology Department of the University of L'Aquila (Avezzano).

Data will be collected before mAbs starting (baseline-T0) and after three months (T1) of mAbs treatment. First, Repsonder and Non-responder groups will be identified, then a biochemical profiling of the two subgroups will be performed at T0 and T1.

METHODS:

All patients will undergo a biochemical profiling that will include analysis of:

* eCBome system: mRNS levels of FAAH, MAGL, DAGL, NAPE, NAAA in peripheral blood mononuclear cells,
* plasma levels of AEA, 2-AG, PEA and OEA; CGRP, PACAP, and VIP; IL-1beta, TNF-alpha, IL-4, and IL-10; kynurenic and quinolinic acids;
* miR-382-5p, miR-34a, miR-30a, and miR-155 in peripheral blood mononuclear cells,
* shotgun analysis of microbiota in patients' faeces.

Biochemical sampling will be collected between 9 and 11 a.m. to avoid circadian rhythm influence. All evaluation will be performed in migraine interictal phase.

The following collection methods will be adopted:

* mRNA and microRNA analysis in PBMCs. Blood samples will be collected within ethylenediamine tetra-acetic acid tubes, with a first isolation of PBMCs and total RNA. Ubiquitin C and U6 will act as housekeeping genes for genes coding for the eCBome enzymes and miRNAs.
* kynurenic acid and quinolinic acid, AEA, PEA, 2-AG and OEA will be determined according to Gao published method (Gao, 2020). Kinurenine metabolite levels will be measured according to the method described by Fuertig (Fuertig, 2016).
* CGRP alpha, PACAP-38 and VIP levels will be measured using a commercial enzyme linked immunosorbent assay
* IL-1beta, TNF-alpha, IL-4, IL-10 cytokine will be measured by the Ella Automated Immunoassay System with a Simple Plex assay panel.
* Microbiome analysis: after correct collection and preservation of stool specimens, they will be delivered to the Translational Neurovascular Research Unit (IRCCS Mondino Foundation) for DNA extraction.

STATISTICAL ANALYSIS Sample size calculation is defined for primary outcome (MAGL expression), while a power analysis is performed for the co-primary outcome (FAAH expression).

According to preliminary data from the work of Greco 2021 suggesting a ratio between Non-responders and Responders: 2:3 and MAGL gene expression: 8±10 RQ in Non-responders and 3±4 RQ in Responders, the minimum sample size is of 88 migraine patients (53 Responders and 35 NON-Responders) in order to have a confidence interval 95% and power of 80%.

Normality analysis will be performed to evaluate parametric or non-parametric methods.

A univariate analysis will be performed to search for differences in demographic, clinical and biochemical parameters between Non-Responder and Responder groups at T0. Main statistical analysis will include a multivariate approach to control for confounders. The level of significance will be set at alpha = 0.05 considering correction for multiple comparisons where appropriate.

Eligibility criteria: Inclusion Criteria:

* male and female patients aged 18 to 75 years
* diagnosis of episodic migraine or chronic migraine according to ICHD-3 criteria
* for episodic migraine: 8-14 monthly migraine days in the previous 3 months
* diagnosis of resistant migraine defined by: i) having failed at least 3 classes of migraine preventatives and ii) suffering from at least 8 debilitating monthly headache days for at least 3 consecutive months
* patients naive to CGRP targeting treatments

Exclusion Criteria:

* history of major psychiatric or other neurological conditions
* diagnosis of other primary or secondary headache disorders (only sporadic tension-type headache is allowed if the patients can clearly differentiate between the 2 types of headaches)
* changes in ongoing preventive treatment (if any) in the previous 3 months
* clinically significant medical conditions
* chronic pain conditions
* alcohol and/or drug abuse
* pregnancy or lactation

Primary outcomes:
measure: Gene expression of FAAH
timeFrame: Baseline (T0) - three months of mAbs treatment (T1)
measure: Gene expression of MAGL
timeFrame: Baseline (T0) - three months of mAbs treatment (T1)
Secondary outcomes:
measure: Gene expression of catalyzing enzymes (DAGL, NAPE, NAAA)
timeFrame: Baseline (T0) - three months of mAbs treatment (T1)
measure: Plasma levels of AEA, 2-AG, PEA, OEA
timeFrame: Baseline (T0) - three months of mAbs treatment (T1)
measure: Plasma levels of CGRP, PACAP and VIP
timeFrame: Baseline (T0) - three months of mAbs treatment (T1)
measure: Gene expression of miR-382-5p, miR-34a, miR-30a and miR-155
timeFrame: Baseline (T0) - three months of mAbs treatment (T1)
measure: Plasma levels of IL-1beta, TNF-alpha, IL-4 and IL-10
timeFrame: Baseline (T0) - three months of mAbs treatment (T1)
measure: Plasma levels of kynurenic acid and quinolinic acid
timeFrame: Baseline (T0) - three months of mAbs treatment (T1)
measure: Shotgun analysis of microbiota
timeFrame: Baseline (T0) - three months of mAbs treatment (T1)
Other outcomes:
Locations:
city: Pavia
country: Italy
facility: Headache Science & Neurorehabilitation Unit
Study officials:
References:
citation: Gao W, Walther A, Wekenborg M, Penz M, Kirschbaum C. Determination of endocannabinoids and N-acylethanolamines in human hair with LC-MS/MS and their relation to symptoms of depression, burnout, and anxiety. Talanta. 2020 Sep 1;217:121006. doi: 10.1016/j.talanta.2020.121006. Epub 2020 Apr 9.
citation: Fuertig R, Ceci A, Camus SM, Bezard E, Luippold AH, Hengerer B. LC-MS/MS-based quantification of kynurenine metabolites, tryptophan, monoamines and neopterin in plasma, cerebrospinal fluid and brain. Bioanalysis. 2016 Sep;8(18):1903-17. doi: 10.4155/bio-2016-0111. Epub 2016 Aug 15.
citation: Greco R, Demartini C, Zanaboni AM, Tumelero E, Icco R, Sances G, Allena M, Tassorelli C. Peripheral changes of endocannabinoid system components in episodic and chronic migraine patients: A pilot study. Cephalalgia. 2021 Feb;41(2):185-196. doi: 10.1177/0333102420949201. Epub 2020 Sep 23.
citation: De Icco R, Greco R, Demartini C, Vergobbi P, Zanaboni A, Tumelero E, Reggiani A, Realini N, Sances G, Grillo V, Allena M, Tassorelli C. Spinal nociceptive sensitization and plasma palmitoylethanolamide levels during experimentally induced migraine attacks. Pain. 2021 Sep 1;162(9):2376-2385. doi: 10.1097/j.pain.0000000000002223.