Dissertation committee:
Μαρία Αναγνωστούλη, Επίκουρη Καθηγήτρια, Ιατρική Σχολή, ΕΚΠΑ
Γεώργιος Χρούσος, Ομότιμος Καθηγητής, Ιατρική Σχολή, ΕΚΠΑ
Μαρία Ροζέ Ροντρίγκεθ Πονς, Καθηγήτρια, Ιατρική Σχολή, ΕΚΠΑ
Λεωνίδας Στεφανής,Καθηγητής, Καθηγητής, Ιατρική Σχολή, ΕΚΠΑ
Γεώργιος Κούτσης, Αναπληρωτής Καθηγητής, Ιατρική Σχολή, ΕΚΠΑ
Μαρία Ελευθερία Ευαγγελοπούλου, Αναπληρώτρια Καθηγήτρια, Ιατρική Σχολή, ΕΚΠΑ
Αργύριος Ντινόπουλος, Καθηγητής Ιατρική Σχολή, ΕΚΠΑ
Summary:
Introduction: Pediatric-onset multiple sclerosis (POMS) accounts approximately for 3-5% of all cases of MS. In 2013, the International Pediatric Multiple Sclerosis Study Group (IPMSSG) established diagnostic criteria for all acquired childhood demyelinating diseases. Since then, the global scientific community has focused more intensely on investigating MS, examining both the clinical and imaging features of the disease, which highlight a highly inflammatory and possibly less degenerative profile, as well as the genetic and immunological mechanisms that contribute to its manifestation. The association of the Major Histocompatibility Complex (MHC) and specifically the HLA-DRB1*15:01 allele with both pediatric and adult disease is now commonly accepted. Furthermore, while previous studies have reported that Apolipoprotein E (ApoE) gene polymorphisms are not associated with MS, ApoE2, and ApoE4 alleles have been repeatedly associated with depression, sleep disorders, cognitive problems, and stress in the context of MS, while data on the role of ApoE in POMS are absent from the literature.
Aim of the study: The present study aimed to characterize the genotypes of HLADRB1* and ApoE gene polymorphisms in an expanded cohort of POMS patients. Furthermore clinical, imaging, and laboratory characteristics of these patients as well as parameters related to sleep quality and stress have been evaluated. Secondarily, our study aimed to highlight any differences in these parameters compared to adult-onset MS (AOMS) patients and the general population, as well as to identify possible associations of HLADRB1* and ApoE alleles and genotypes with these parameters.
Materials and Methods: One hundred (100) patients with disease onset of ≤19 years of age, fulfilling the revised IPMSSG 2013 criteria were included in the study, enrolled both prospectively and retrospectively, and for which demographic, clinical, imaging and laboratory data were recorded. For all patients, written informed consent was signed by themselves or by their legal guardian. The results were compared with those of 168 AOMS patients. HLA-DRB1 and ApoE genotyping was performed with a low-resolution SSO technique. The observed allele frequencies were compared with those of a Greek healthy control’s cohort (n=246) genotyped in a previous study. ApoE allele frequencies were compared with those of 391 AOMS patients and 200 healthy controls genotyped in a previous study, including a number of patients of our Laboratory. Forty (40) patients consented to answer the questionnaire to assess sleep quality. Statistical processing was performed with SPSS, SPSS Inc, Chicago, IL, USA. Univariate comparisons were performed using Mann-Whitney U and chi-square tests (with Yates correction for 2x2 tables) for intervals and categorical variables, respectively. Hardy-Weinberg ratios for HLA-DRB1 and ApoE haplotypes were confirmed using PyPoP software, and homozygosity/heterozygosity neutrality testing was performed using Ewens-Watterson tests. The statistical significance level was set at α=0.05. In tests related to HLA genotype (excluding clinical parameters), p-value correction was performed using the Benjamini-Yekutieli (or B-Y) method (p≤ 0.016).
Results: The mean age of disease onset was estimated at 15.5 ± 2.5 years and the incidence was more frequent in women at 61%. POMS patients had significantly lower EDSS [mean±SD: 2.5±1.43 vs 3.3±1.7, p <0.001] compared to AOMS patients. POMS patients display significantly higher frequencies of the HLA-DRB1*15 allele (35% vs. 16.7%, OR [95%CI]: 2.69 (1.58 -4.58), p =0.001) compared to controls. Significantly increased prevalence of HLA-DRB1*03 was also observed in POMS compared to both healthy controls (24% vs. 12.6%, OR [95%CI]: 2.19 (1.21-3.97), p=0.016) and AOMS (24% vs. 13.1%, OR [95%CI]: 2.1 (1.1-3.98), p=0.034, but not significant following B-Y correction). In addition, HLA-DRB1*03-positive patients display reduced risk for brainstem lesions development (OR [CI 95%]:0.19 (0.06-0.65), p=0.011). Additionally, a significantly lower frequency of HLA-DRB01*07 (4% vs 13.4%, OR (95% CI): 0.27 (0.09-0.78), p= 0.017* but marginally not significant following B-Y correction) and HLA-DRB01*11 (37% vs 52%, OR [95% CI]: 0.54 (0.34-0.87), p= 0.016) was observed in POMS patients compared to healthy controls.
Regarding ApoE genotyping, a significantly higher incidence of ApoE2/E3 and E3/E4 genotypes in POMS patients compared to the general population (20.5% vs 11%, OR [95%]: 2.1 (1.1-4.0), p = 0.02) and (17.9% vs 10%, OR [95%]: 1.95 (1.1-3.4), p = 0.05) respectively) was observed. Similarly, for the first time, a significantly higher frequency of ApoE2 and ApoE4 (marginally significant) alleles was observed in POMS patients compared to healthy controls (11% vs 5.3%, OR [95%]: 2.45 (1.3-4.62), p=0.008 and 9.8% vs 5.8%, OR [95%]: 2.1 (1.1-3.99), p=0.045 respectively). Moreover, a significantly lower frequency of the ApoE3 allele was found in POMS patients compared to both healthy controls (79% vs 89%, OR [95%]: 0.46 (0.30-0.72), p=0.0007) and AOMS (79% vs 85.4%, OR [95%]: 0.62 (0.43-0.90), p=0.014). Similarly, a significantly lower frequency of the ApoE3/E3 genotype was detected in POMS patients compared to both healthy controls (59.8% vs 79%, OR [95%]: 0.40 (0.24-0.25), p=0.0003) and AOMS (59.8% vs 71.6%, OR [95%]: 0.6 (0.40-0.90), p=0.02).
Finally, regarding sleep quality, among the 40 POMS patients who consented to answer the questionnaire, the most frequent disorder was interrupted sleep (62.5%), followed by hyperactivity/anxiety disorder (25%) and delayed sleep onset (12.5%).
Conclusions: POMS patients display, a lower EDSS score compared to AOMS. Regarding immunogenetic findings, the HLA-DRB1*15 and HLA-DRB1*03 alleles emerge as the main risk factors for disease development compared to AOMS, indicating the genetic homogeneity of the disease between the two clinical subtypes, concerning HLA-DRB1*15 allele. Similarly, to NMO, the HLA-DRB1*03 allele was associated with an increased risk of developing POMS, as well as a higher risk for brainstem lesions development, indicating a possible increased role of humoral immunity in POMS patients. Regarding ApoE polymorphisms, the ApoE2 allele together with the ApoE2/E3 genotype seems to be a significant risk factor for POMS development in our POMS cohort. Finally, regarding the sleep quality and stress, interrupted sleep seems to be the most common disorder among POMS patients with sleep disorders. However, this finding should be confirmed in a larger patient cohort.
Further investigation of the immunogenetic differences between POMS and AOMS regarding the genetic region of HLADRB1* alleles, and the role of ApoE in a larger number of patients is required. The investigation of the role of these two aforementioned genetic loci, using -omics methods such as transcriptomics and proteomics could potentially highlight new distinct pathophysiological mechanisms between POMS and AOMS, with reflection on therapeutic intervention as well.
