NGS-Based Identification of Two Novel PCDH19 Mutations in Female Patients with Early-Onset Epilepsy.

Developmental and epileptic encephalopathy-9 (DEE9) is characterized by seizure onset in infancy, mild to severe intellectual impairment, and psychiatric features and is caused by a mutation in the PCDH19 gene on chromosome Xq22. The rare, unusual X-linked type of disorder affects heterozygous females and mosaic males; transmitting males are unaffected. In our study, 165 patients with epilepsy were tested by Next Generation Sequencing (NGS)-based panel and exome sequencing using Illumina technology. PCDH19 screening identified three point mutations, one indel, and one 29 bp-long deletion in five unrelated female probands. Two novel mutations, c.1152_1180del (p.Gln385Serfs*6) and c.830_831delinsAA (p.Phe277*), were identified and found to be de novo pathogenic. Moreover, among the three inherited mutations, two originated from asymptomatic mothers and one from an affected father. The PCDH19 c.1682C>T and c.1711G>T mutations were present in the DNA samples of asymptomatic mothers. After targeted parental testing, X chromosome inactivation tests and Sanger sequencing were carried out for mosaicism examination on maternal saliva samples in the two asymptomatic PCDH19 mutation carrier subjects. Tissue mosaicism and X-inactivation tests were negative. Our results support the opportunity for reduced penetrance in DEE9 and contribute to expanding the genotype-phenotype spectrum of PCDH19-related epilepsy.

Importance and application of WES in fetal genetic diagnostics: Identification of novel ASPM mutation in a fetus with microcephaly.

Background: Prenatal whole exome sequencing (WES) approaches can provide genetic diagnosis with rapid turnaround time and high diagnostic rate when conventional tests are negative. Here we report a family with multiple pregnancy loss and with repeated occurrence of fetal microcephaly. Methods and results: Because of positive family history and recurrent structural abnormality during the pregnancies that may lead postnatal neurodevelopmental consequences, WES analysis was indicated. Umbilical cord blood sampling was carried out and WES was performed using Twist Human Core Exome Kit and Illumina sequencing technology. The presence of pathogenic variants was confirmed by Sanger sequencing. WES analysis revealed a known pathogenic c.8506_8507delCA (p.Gln2836Glufs*35, rs587783280) and a novel pathogenic c.3134_3135delTC (p.Leu1045Glnfs*17) ASPM mutations in the fetus in compound heterozygous state. The c.3134_3135delTC has never been reported in the literature. Conclusions: Our findings serve additional evidence that WES can be an efficient and relevant tool to diagnose certain genetic disorders with appropriate indication and to assess the recurrence risk of a disease. With the application of WES in combination with pre-implantation genetic tests, we can avoid the transmission of pathogenic mutations and we can achieve a decreased abortion rate in obstetric care.

Overlapping Interstitial Deletions of the Region 9q22.33 to 9q33.3 of Three Patients Allow Pinpointing Candidate Genes for Epilepsy and Cleft Lip and Palate.

Introduction: Patients carrying interstitial deletions of the long arm of chromosome 9 show similar features. These phenotypes are often characterized by developmental delay, intellectual disability, short stature, and dysmorphism. Previously reported deletions differ in size and location spanning from 9q21 to 9q34 and were mostly detected by conventional cytogenetic techniques. Methods: Based on clinical features suggesting primarily chromosomal diseases, aCGH analysis was indicated. We report on de novo overlapping interstitial 9q deletions in 3 unrelated individuals presenting neurodevelopmental disorder and multiple congenital anomalies. Results: An 8.03-Mb (90 genes), a 15.71-Mb (193 genes), and a 15.81-Mb (203 genes) deletion were identified in 9q affecting 9q22.33q33.3. The overlapping region was 1.50 Mb, including 2 dosage-sensitive genes, namely EPB41L4B (OMIM #610340) and SVEP1 (OMIM #611691). These genes are thought to be involved in cellular adhesion, migration, and motility. The non-overlapping regions contain 24 dosage-sensitive genes. Conclusion: Besides the frequently described symptoms (developmental delay, intellectual disability, skeletal abnormalities, short stature, and dysmorphic facial features) shared by the patients with interstitial deletions of chromosome 9q reported thus far, two of our patients showed distinct forms of epilepsy, which were successfully treated, and one had a bilateral cleft lip and palate. Possible candidate genes for epilepsy and cleft lip and palate are discussed.

Correlation between large FBN1 deletions and severe cardiovascular phenotype in Marfan syndrome: Analysis of two novel cases and analytical review of the literature.

BACKGROUND: Marfan syndrome (MFS) is a clinically heterogeneous hereditary connective tissue disorder. Severe cardiovascular manifestations (i.e., aortic aneurysm and dissection) are the most life-threatening complications. Most of the cases are caused by mutations, a minor group of which are copy number variations (CNV), in the FBN1 gene. METHODS: Multiplex ligation-dependent probe amplification test was performed to detect CNVs in 41 MFS patients not carrying disease-causing mutations in FBN1 gene. Moreover, the association was analyzed between the localization of CNVs, the affected regulatory elements and the cardiovascular phenotypes among all cases known from the literature. RESULTS: A large two-exon deletion (exon 46 and 47) was identified in two related patients, which was associated with a mild form of cardiovascular phenotype. Severe cardiovascular symptoms were found significantly more frequent in patients with FBN1 large deletion compared to our patients with intragenic small scale FBN1 mutation. Bioinformatic data analyses of regulatory elements located within the FBN1 gene revealed an association between the deletion of STAT3 transcription factor-binding site and cardiovascular symptoms in five out of 25 patients. CONCLUSION: Our study demonstrated that large CNVs are often associated with severe cardiovascular manifestations in MFS and the localization of these CNVs affect the phenotype severity.

Genotype-Phenotype Associations in Patients With Type-1, Type-2, and Atypical NF1 Microdeletions.

Neurofibromatosis type 1 is a tumor predisposition syndrome inherited in autosomal dominant manner. Besides the intragenic loss-of-function mutations in NF1 gene, large deletions encompassing the NF1 gene and its flanking regions are responsible for the development of the variable clinical phenotype. These large deletions titled as NF1 microdeletions lead to a more severe clinical phenotype than those observed in patients with intragenic NF1 mutations. Around 5-10% of the cases harbor large deletion and four major types of NF1 microdeletions (type 1, 2, 3 and atypical) have been identified so far. They are distinguishable in term of their size and the location of the breakpoints, by the frequency of somatic mosaicism with normal cells not harboring the deletion and by the number of the affected genes within the deleted region. In our study genotype-phenotype analyses have been performed in 17 mostly pediatric patients with NF1 microdeletion syndrome identified by multiplex ligation-dependent probe amplification after systematic sequencing of the NF1 gene. Confirmation and classification of the NF1 large deletions were performed using array comparative genomic hybridization, where it was feasible. In our patient cohort 70% of the patients possess type-1 deletion, one patient harbors type-2 deletion and 23% of our cases have atypical NF1 deletion. All the atypical deletions identified in this study proved to be novel. One patient with atypical deletion displayed mosaicism. In our study NF1 microdeletion patients presented dysmorphic facial features, macrocephaly, large hands and feet, delayed cognitive development and/or learning difficulties, speech difficulties, overgrowth more often than patients with intragenic NF1 mutations. Moreover, neurobehavior problems, macrocephaly and overgrowth were less frequent in atypical cases compared to type-1 deletion. Proper diagnosis is challenging in certain patients since several clinical manifestations show age-dependency. Large tumor load exhibited more frequently in this type of disorder, therefore better understanding of genotype-phenotype correlations and progress of the disease is essential for individuals suffering from neurofibromatosis to improve the quality of their life. Our study presented additional clinical data related to NF1 microdeletion patients especially for pediatric cases and it contributes to the better understanding of this type of disorder.

Maternal mosaicism underlies the inheritance of a rare germline AKT3 variant which is responsible for megalencephaly-polymicrogyria-polydactyly-hydrocephalus syndrome in two Roma half-siblings.

Megalencephaly-polymicrogyria-polydactyly-hydrocephalus (MPPH) syndrome is a developmental brain disorder characterized by an enlarged brain size with bilateral perisylvian polymicrogyria and a variable degree of ventriculomegaly. MPPH syndrome is associated with oromotor dysfunction, epilepsy, intellectual disability and postaxial hexadactyly. The molecular diagnosis of this disorder is established by the identification of a pathogenic variant in either AKT3, CCND2 or PIK3R2. Previously reported AKT3 variants are associated with various brain abnormalities and may lead to megalencephaly. MPPH syndrome is usually due to germline pathogenic AKT3 variants. Somatic mosaic pathogenic variants associated with hemimegalencephaly, which is similar to MPPH, have also been observed. A Hungarian Roma family with two half-siblings, which present with intellectual disability, dysmorphic features, epilepsy, brain malformations, and megalencephaly was studied. Whole exome sequencing (WES) analysis was performed. WES analysis revealed a heterozygous c.1393C > T p.(Arg465Trp) pathogenic missense AKT3 variant in both affected half-siblings. The variant was verified via Sanger sequencing and was not present in the DNA sample from the healthy mother, which was derived from peripheral blood, suggesting maternal germline mosaicism. In conclusion, this is the first report in which maternal germline mosaicism of a rare pathogenic AKT3 variant leads to autosomal dominantly inherited MPPH syndrome.

[A rare form of ion channel gene mutation identified as underlying cause of generalized epilepsy]. / Generalizált epilepszia hátterében azonosított ioncsatorna-génmutáció ritka formája.

The advances in molecular genetic methods has lead to the discovery of the genetic alterations that underlie the etiology of most diseases previously held to be idiopathic. Targeted genetic examination of a pediatric male patient showing a normal intellect, an extended area of skin hypopigmentation, and suffering from generalized epilepsy displaying a switch in epilepsy syndrome during the course of the disease towards a neurocutaneous syndrome was unsuccessful. Whole-exome sequencing identified a heterozygous missense mutation in a potassium chloride cotransporter gene, which together with the phenotype underscores the diagnosis of an epilepsy syndrome known in the literature as idiopathic generalized epilepsy type 14. Orv Hetil. 2019; 160(21): 835-838.

Age-Related Hearing Impairment Associated NAT2, GRM7, GRHL2 Susceptibility Gene Polymorphisms and Haplotypes in Roma and Hungarian Populations.

Age-related hearing impairment (ARHI) is the most frequent sensory disease in the elderly, which is caused by an interaction between genetic and environmental factors. Here we examined the ethnic differences, allele and genotype frequencies of the NAT2, GRM7, and GRHL2 genes pooled samples of healthy Hungarian and healthy and hearing impaired Roma people. Study populations of healthy Hungarian and Roma subjects were characterized for the rs1799930 NAT2, rs11928865 GRM7, rs10955255, rs13263539, and rs1981361 GRHL2 polymorphisms and deaf Roma subjects were characterized for the rs1799930 NAT2, rs13263539, and rs1981361 GRHL2 using a PCR-RFLP method. We found significant differences in minor allele frequencies for GRHL2 rs13263539 and rs1981361 polymorphism between healthy Roma and Hungarian samples (37.9% vs. 51.0% and 43.6% vs. 56.2%, respectively; p < 0.05). The differences of homozygous genotype of GRHL2 rs13263539 and rs1981361 variants, values were also significantly different (13.0% vs. 25.3% and 16.5 vs. 32.3%; p < 0.05). The NAT2 rs1799930 homozygous genotype was 14.0% in healthy Romas and 7.7% in Hungarians, while the minor A allele frequency was 38.0% and 26.7% in Roma and Hungarian population, respectively (p < 0.05). Furthermore, the frequency of GGT, GAC and GAT haplotypes was significantly higher in the Hungarian population than in healthy Roma (1.87 vs. 4.47%, 0.91 vs. 2.07% and 1.15 vs. 5.51%, respectively; p < 0.008). Present study revealed significant interethnic differences in allele polymorphisms of NAT2, GRM7 and GRHL2 exhibit quite marked ethnic differences in Roma populations that might have important implications for the preventive and therapeutic treatments in this population.

Cytochrome P450 Drug Metabolizing Enzymes in Roma Population Samples: Systematic Review of the Literature.

The cytochrome P450 drug metabolizing enzymes are highly polymorphic and show inter-individual differences in variability in drug response, which varies widely also with ethnicity. This study aims to summarize the available data on genetic polymorphisms associated with cytochrome enzymes conducted on Roma populations. Our goal was to compare the frequency of the variant alleles, genotypes and predicted phenotypes with corresponding rates from other populations. We carried out a systematic review including the papers published on the pharmacogenetically relevant variants of cytochrome P450 genes related to Roma population. The study was performed using several articles, websites and databases, including PubMed, Ensembl, dbSNP, HapMap and 1000 Genomes Project. This review attempts to summarize and discuss our current knowledge about the frequency distribution of the ever investigated 20 allelic variants of 9 cytochrome genes (CYP1A2, CYP1B1, CYP2B6, CYP2C9, CYP2C19, CYP2C8, CYP2D6, CYP3A5, CYP4F2) in Roma DNA samples and compare them with other populations. Differences between Roma and Hungarian samples are reported for 7 variant genotypes. CYP2C9 *2/*3 and CYP2C19 *2/*2 genotypes showed more than 3-fold differences. Additional differences are displayed for allele frequency of 7 variants (rs762551, rs3745274, rs1058930, rs1065852, rs3892097, rs1057910 and rs4244285) in Roma population samples. The interethnic variability in clinically relevant genetic polymorphisms of drug metabolizing enzymes, which may explain distinct drug response, highlights the need to allow for the ancestry of participants in pharmacogenetic studies.

Kleefstra syndrome in Hungarian patients: additional symptoms besides the classic phenotype.

BACKGROUND: Kleefstra syndrome is a rare genetic disorder, with core phenotypic features encompassing developmental delay/intellectual disability, characteristic facial features - brachy(micro)cephaly, unusual shaped eyebrows, flat face with hypertelorism, short nose with anteverted nostrils, thickened lower lip, carpmouth with macroglossia - and childhood hypotonia. Some additional symptoms are observed in different percentage of the patients. Epilepsy is common symptom as well. The underlying cause of the syndrome is a submicroscopic deletion in the chromosomal region 9q34.3 or disruption of the euchromatin histone methyl transferase 1. CASE PRESENTATION: We describe two Hungarian Kleefstra syndrome patients, one with the classic phenotype of the syndrome, the diagnosis was confirmed by subtelomeric FISH. Meanwhile in our second patient beside the classic phenotype a new symptom - abnormal antiepileptic drug metabolic response - could be observed. Subtelomere FISH confirmed the 9q34.3 terminal deletion. Because of the abnormal drug metabolism in our second patient, we performed array CGH analysis as well searching for other rearrangements. Array CGH analysis indicated a large - 1.211 Mb -, deletion only in the 9q subtelomeric region with breakpoints ch9:139,641,471-140,852,911. CONCLUSIONS: This is the first report on Kleefstra syndrome in patients describing a classical and a complex phenotype involving altered drug metabolism.

Interethnic differences of cytochrome P450 gene polymorphisms may influence outcome of taxane therapy in Roma and Hungarian populations.

Taxanes are widely used microtubule-stabilizing chemotherapeutic agents in the treatment of cancers. Several cytochrome P450 gene variants have been proven to influence taxane metabolism and therapy. The purpose of this work was to determine the distribution of genetic variations of CYP1B1, CYP2C8 and CYP3A5 genes as the first report on taxane metabolizer cytochrome P450 gene polymorphisms in Roma and Hungarian populations. A total of 397 Roma and 412 Hungarian healthy subjects were genotyped for CYP1B1 c.4326C > G, CYP2C8 c.792C > G and CYP3A5 c.6986A > G variant alleles by PCR-RFLP assay and direct sequencing. We found significant differences in the frequencies of homozygous variant genotypes of CYP1B1 4326 GG (p = 0.002) and CYP3A5 6986 GG (p < 0.001) between Roma and Hungarian populations. Regarding minor allele frequencies, for CYP2C8 a significantly increased prevalence was found in 792G allele frequency in the Hungarian population compared to the Roma population (5.83% vs. 2.14%, p = 0.001). Our results can be used as possible predictive factors in population specific treatment algorithms to developing effective programs for a better outcome in patients treated with taxanes.

Marked differences in frequencies of statin therapy relevant SLCO1B1 variants and haplotypes between Roma and Hungarian populations.

BACKGROUND: SLCO1B1 polymorphisms are relevant in statin pharmacokinetics. Aim of this study was to investigate the genetic variability and haplotype profile of SLCO1B1 polymorphisms in Roma and Hungarian populations. Genotypes of 470 Roma and 442 Hungarian subjects for c.388A > G, c.521T > C and c.1498-1331T > C polymorphisms were determined by PCR-RFLP assay. Using these SNPs eight different haplotypes could be differentiated. RESULTS: Differences were found between Roma and Hungarians in SLCO1B1 388AA (24.5 vs. 45.5 %), GG (33.4 vs. 17.9 %) genotypes, AG + GG (75.5 vs. 54.5 %) carriers, in G allele frequency (0.545 vs. 0.362), respectively (p < 0.001). The most common SLCO1B1 haplotype was the ht8 (GTT) both in Roma (43.6 %) and in Hungarian (59.1 %) samples. The ht6 (GCT) was not present in Roma population samples Haplotype analyses showed striking differences between the Roma and Hungarian samples in ht4 (ATT, 37.2 % vs 20.8 %), ht5 (GCC, 1.15 % vs. 3.62 %) and ht8 (GTT, 43.6 % vs. 59.1 %) haplotypes (p < 0.01), respectively. Linkage disequilibrium analysis showed that the studied variants are in different linkage disequilibrium patterns depending on the ethnic origin. CONCLUSIONS: Similarly to Caucasians the 388G is the minor allele in Hungarians, however, in Roma the 388A was found to be the minor allele contrary to Indians (India). The minor allele frequency of 521T > C and 1498-1331T > C SNPs are almost three times higher in Romas than in Indians (Singapore and Gujarati, respectively). Observed allele frequency for 1498-1331T > C polymorphism reflects the measured average European rates in Hungarians. The results can be applied in population specific treatment algorithms when developing effective programs for statin therapy.

Interethnic variability of CYP4F2 (V433M) in admixed population of Roma and Hungarians.

AIMS: Pharmacogenetic based dosing recommendations are provided in FDA-approved warfarin label for Caucasians. Evidence of notable difference in dosing algorithms of under-represented populations forced us to explore the genetic variability of CYP4F2 gene in Roma and Hungarian populations. PATIENTS AND METHODS: 484 Roma, 493 Hungarian untreated subjects were genotyped for the CYP4F2*3 (rs2108622) variant by PCR-RFLP assay. RESULTS AND DISCUSSION: We firstly report, that frequencies of the CYP4F2 rs2108622 GG, GA, AA genotypes and A allele in the Roma population were 46.5%, 42.6%, 10.9% and 32.2%; in Hungarians 50.1%, 42.2%, 7.7% and 22.8%, respectively. Bearing of two minor alleles of CYP4F2 missense variant (AA genotype) modestly explains inter-ethnic differences of studied populations (p<0.08). CYP4F2*3 (V433M) risk allele frequency of Roma (0.32) was in higher range, and of Hungarians (0.23) in lower range, as compared with other world populations. CONCLUSIONS: Roma have an elevated chance for higher mean warfarin dose, besides a decreased risk of major bleeding events in long-term warfarin use.

Increased prevalence of functional minor allele variants of drug metabolizing CYP2B6 and CYP2D6 genes in Roma population samples.

BACKGROUND: Cytochrome P450 2B6 and 2D6 are important enzymes in human drug metabolism. These phase I enzymes are known to contribute the biotransformation of clinically important pharmaceuticals, including antidepressants, anticancer and anxiolytic drugs. The aim of this work was to determine the pharmacogenetic profile of CYP2B6 and CYP2D6 in Roma and Hungarian population samples. METHODS: A study population of 426 healthy Roma and 431 healthy Hungarian subjects were characterized for CYP2B6 c.516G>T, CYP2D6 c.100C>T and c.1846G>A polymorphisms using predesigned TaqMan Drug Metabolism Genotyping Real Time-PCR assays. RESULTS: We found significant differences in the presence of CYP2B6 c.516G>T (p<0.001), CYP2D6 c.100C>T (p=0.003) and c.1846G>A (p=0.022) between Hungarian and Roma population. The 516T allele frequency was 33.6% in the Roma group, 21.4% in Hungarians, whereas the minor CYP2D6 100T allele was present in 26.6% in Romas and 20.5% in Hungarians. The 1864A allele frequency was 22.5% in Roma and 18.1% in Hungarian samples. A significant increase was found in genotype frequencies for homozygous minor allele carrier Roma participants compared to Hungarians for CYP2B6 516TT and CYP2D6 100TT. The following CYP2D6 genotypes were identified in Roma samples: *1/*1 (55.4%), *1/*4 (2.1%), *1/*10 (3.1%), *4/*10 (38.7%), *10/*10 (0.7%). CONCLUSION: Our results demonstrate an increased minor allele frequency for CYP2B6 and CYP2D6 polymorphisms in Roma samples that implies clinical significance in this ethnic group.

Extreme differences in SLCO1B3 functional polymorphisms in Roma and Hungarian populations.

Variants in SLCO1B3 transporter are linked to disposition and uptake of drugs and show high degree of heterogeneity between populations. A total of 467 Roma and 448 Hungarian subjects were genotyped for SLCO1B3 c.334T>G and c.1683-5676A>G variant alleles by PCR-RFLP assay and direct sequencing. We found significant differences in the frequencies of homozygous variant genotypes of SLCO1B3 334GG (41.54% vs. 8.04%, p<0.001) and 1683-5676GG (0.43% vs. 2.01%, p=0.028) between Romas and Hungarians. A significantly increased prevalence was found in SLCO1B3 1683-5676G allele frequency in Hungarians compared to the Roma population (15.07% vs. 3.43%, p≤0.001). The frequency of SLCO1B3 334G allele was significantly increased in Roma population compared to Hungarians (70.56% vs. 52.23%, p=0.001). The LD values between the examined SNPs were 80 and 90 in Roma and in Hungarian samples, respectively. Our results highlight notable pharmacogenetic differences between Roma and Hungarian populations, which may have therapeutic implications.

Functional variants of lipid level modifier MLXIPL, GCKR, GALNT2, CILP2, ANGPTL3 and TRIB1 genes in healthy Roma and Hungarian populations.

The role of triglyceride metabolism in different diseases, such as cardiovascular or cerebrovascular diseases is still under extensive investigations. In genome-wide studies several polymorphisms have been reported, which are highly associated with plasma lipid level changes. Our goal was to examine eight variants: rs12130333 at the ANGPTL3, rs16996148 at the CILP2, rs17321515 at the TRIB1, rs17145738 and rs3812316 of the MLXIPL, rs4846914 at GALNT2, rs1260326 and rs780094 residing at the GCKR loci. A total of 399 Roma (Gypsy) and 404 Hungarian population samples were genotyped using PCR-RFLP method. Significant differences were found between Roma and Hungarian population samples in both MLXIPL variants (C allele frequency of rs17145738: 94.1% vs. 85.6%, C allele frequency of rs3812316: 94.2% vs. 86.8% in Romas vs. in Hungarians, p < 0.05), in ANGPTL3 (T allele frequency of rs1213033: 12.2% vs. 18.5% in Romas vs. Hungarians, p < 0.05) and GALNT2 (G allele frequency of rs4846914: 46.6% vs. 54.5% Romas vs. in Hungarians, p < 0.05), while no differences over SNPs could be verified and the known minor alleles showed no correlation with triglyceride levels in any population samples. The current study revealed fundamental differences of known triglyceride modifying SNPs in Roma population. Failure of finding evidence for affected triglyceride metabolism shows that these susceptibility genes are much less effective compared for example to the apolipoprotein A5 gene.

Genetic polymorphisms in promoter and intronic regions of CYP1A2 gene in Roma and Hungarian population samples.

The purpose of this study was to determine the interethnic differences of four CYP1A2 drug metabolizing enzyme variants. A total of 404 Roma and 396 Hungarian healthy subjects were genotyped for -163C>A, -729C>T, -2467delT and -3860G>A variants of CYP1A2 by RT-PCR and PCR-RFLP technique. The -3860A and -729T allele were not detectable in Roma samples, while in Hungarian samples were present with 2.02% and 0.25% prevalence, respectively. There was a 1.5-fold difference in presence of homozygous -163AA genotype between Hungarian and Roma samples (49.5% vs. 31.9%, p<0.001). The -163A allele frequency was 68.6% in Hungarians and 56.9% in Romas (p=0.025). The -2467delT allele frequency was 6.81% in Roma group and 5.81% in Hungarians. The most frequent allelic constellation was -3860G/-2467T/-729C/-163A in both populations. In conclusion, Hungarians have markedly elevated chance for rapid metabolism of CYP1A2 substrates, intensified procarcinogen activation and increased risk for cancers.

Admixture of beneficial and unfavourable variants of GLCCI1 and FCER2 in Roma samples can implicate different clinical response to corticosteroids.

Variants of glucocorticoid induced transcript 1 (GLCCI1) result decreased response to inhaled corticosteroids, while intronic variant of low-affinity IgE receptor (FCER2) is associated with exacerbation rates in children with asthma. We examined the ethnic differences, allele and genotype frequencies of two linked single nucleotide polymorphisms (rs37972, rs37973) of GLCCI1 and rs28364072 intronic variant of FCER2 gene in average Roma and Hungarian population. A study population of 474 healthy Roma and 397 Hungarian subjects were characterized for GLCCI1 and FCER2 polymorphisms using real time polymerase chain reaction (PCR) assay and PCR-restriction fragment length polymorphism method. The rs37972 and rs37973 polymorphisms in GLCCI1 were found in 100% linkage disequilibrium both in Romas and in Hungarians. We found significant differences between the two groups regarding both minor allele frequencies (54.5 vs. 43.8%, p ≤ 0.01) and homozygous genotype (31.6 vs. 21.3%, p ≤ 0.01) of GLCCI1. For FCER2 rs28364072 the homozygous variant genotype was present in 2.8% in Romas, while in Hungarians it was 5.8% (p = 0.032). The opposite changes of these two polymorphisms strongly suggest that contrary current belief analyses of GLCCI1 variants are insufficient for personalised glucocorticoid therapies in different populations.

Lower carrier rate of GJB2 W24X ancestral Indian mutation in Roma samples from Hungary: implication for public health intervention.

The purpose of this work was to characterise the W24X mutation of the GJB2 gene in order to provide more representative and geographicaly relevant carrier rates of healthy Roma subisolates and the Hungarian population. 493 Roma and 498 Hungarian healthy subjects were genotyped for the GJB2 c.71G>A (rs104894396, W24X) mutation by PCR-RFLP assay and direct sequencing. This is the first report on GJB2 W24X mutation in geographically subisolated Roma population of Hungary compared to local Hungarians. Comparing the genotype and allele frequencies of GJB2 rs104894396 mutation, significant difference was found in GG (98.4 vs. 99.8 %), GA (1.62 vs. 0.20 %) genotypes and A (0.8 vs. 0.1 %) allele between the Roma and Hungarian populations, respectively (p < 0.02). None of the subjects of Roma and Hungarian samples carried the GJB2 W24X AA genotype. Considerable result of our study, that the proportion of GJB2 W24X GA heterozygotes and the A allele frequency was eight times higher in Roma than in Hungarians. Considering the results, the mutant allele frequency both in Roma (0.8 %) and in Hungarian (0.1 %) populations is lower than expected from previous results, likely reflecting local differentiated subisolates of these populations and a suspected lower risk for GJB2 mutation related deafness. However, the significant difference in GJB2 W24X carrier rates between the Roma and Hungarians may initiate individual diagnostic investigations and effective public health interventions.

Marked differences of haplotype tagging SNP distribution, linkage, and haplotype profile of IL23 receptor gene in Roma and Hungarian population samples.

Polymorphisms of the interleukin-23 receptor (IL23R) gene have been found to play an important role in the development of several autoimmune diseases. We examined five susceptible (rs10889677, rs1004819, rs2201841, rs11805303, rs11209032), one protective (rs7517847) and two neutral variants (rs7530511, rs1884444) of the IL23R gene in pooled DNA of healthy Roma (Gipsy) and Hungarian population samples. Our aim was to determine the genetic variability of the major haplotype tagging polymorphisms, and the haplotype profile of IL23R between the two groups. We analyzed 273 healthy Roma and 253 Hungarian DNA samples using PCR/RFLP assay. Comparing the five susceptible conferring alleles, there were significant increase (p<0.05), while in the protective alleles, there were decrease in the allele frequencies in Roma population (p<0.05). One of the neutral alleles showed increase, the another one did not differ between the two groups. The haplotype analysis of the SNPs revealed fundamentally different association types of SNPs in the two groups; moreover, the frequencies of the various haplotypes also exhibited strong differences, as of ht4 and ht5 haplotypes were significantly higher, whereas the frequencies of ht2 and ht3 haplotypes were significantly lower in the Roma population than in Hungarians (p<0.05). The data presented here show profound differences in the IL23R genetic profiles in the Roma population, that likely has also clinical implications in respect their possible role in the development of certain immunological diseases.