Clinical and Molecular Analysis in Patients with Peutz-Jeghers Syndrome.

Peutz-Jeghers syndrome (PJS) is a rare hereditary disorder linked to increased cancer risk due to specific genetic variants in the STK11 gene. This study aimed to assess disease manifestations, genetic profiles, and genotype-phenotype correlations in PJS patients. Twenty patients from 14 families with PJS who were followed up at our clinic between 2011 and 2021 were included. Genetic susceptibility to hereditary cancers was assess-ed using targeted next-generation sequencing (NGS) and multiplex ligation-dependent probe amplification (MLPA) of the STK11 gene. Clinical data were also collected and analyzed in conjunction with the genetic findings. Initial symptoms appeared around 18.9 years, predominantly abdominal pain and intussusception. Mucocutaneous lesions were found in 85%, and hamartomatous polyps in 90%. Dysplastic polyps were found in 4 patients, with 3 cases of malignancy. Nextgeneration sequencing identified 11 pathogenic and 3 likely pathogenic mutations, including 3 novel STK11 variants (LRG_319: c.598- 8_601del, LRG_319: c.708_718del, and LRG_319: c.146_147del). Next-generation sequencing diagnostic rate was 78.5% (11/14), and the overall diagnostic rate with NGS and MLPA studies was 85.7% (12/14). Patients without STK11 mutations had later symptom onset and potentially lower cancer risk. Truncated mutations are associated with earlier symptoms and elevated cancer risk. This is the first PJS case series in Turkey using the NGS and MLPA methods. It reports 3 novel mutations and emphasizes the genotype-phenotype relationship of PJS. With further studies, the genotype-phenotype relationship of STK11 variants will be better understood.

Investigation of genotype-phenotype and familial features of Turkish dystrophinopathy patients.

Duchenne muscular dystrophy (DMD) and Becker muscular dystrophy (BMD) are X-linked recessive allelic muscle diseases caused by dystrophin gene mutations. Eight hundred thirty-seven patients admitted between 1997 and 2022 were included in the study. Two hundred twenty patients were analyzed by multiplex PCR (mPCR) alone. Five hundred ninety-five patients were investigated by multiplex ligation-dependent probe amplification (MLPA), and 54 patients were examined by sequencing. Deletion was detected in 60% (132/220) of the cases in the mPCR group only and in 58.3% (347/595) of the cases with MLPA analysis. The rates of deletion and duplication were 87.7% and 12.3%, respectively, in the MLPA analysis. Single exon deletions were the most common mutation type. The introns 43-55 (81.8%) and exons 2-21 (13.1%) regions were detected as hot spots in deletions. It was determined that 89% of the mutations were suitable for exon skipping therapy. The reading frame rule did not hold in 7.6% of D/BMD cases (17/224). We detected twenty-five pathogenic/likely pathogenic variants in sequencing, five of which were novel variants. Nonsense mutation was the most common small mutation (44%). 21% of DMD patients were familial. We detected germline mosaicism in four families (4.3%) in the large rearrangement group and one gonosomal mosaicism in a family with a nonsense mutation. This is the largest study examining genotype and phenotype data in Turkish D/BMD families investigated by MLPA analysis. The reading frame hypothesis is not valid in all cases. Sharing the genotype and phenotype characteristics of these cases in the literature will shed light on the molecular structure of DMD and guide gene therapy research. In genetic counseling, carrier screening in the family and possible gonadal mosaicism should be emphasized.

Clinical Heterogeneity in Patients with Long QT Syndrome and Segregation of Single Nucleotide Variants and Clinical Symptoms in 17 Affected Families.

Introduction: Long QT syndrome (LQTS) is a disorder of ventricular myocardial repolarization characterized by a prolonged QT interval on the electrocardiogram. It increases the risk of ventricular arrhythmias, which can cause syncope or sudden cardiac death. In this study, we study the genotype-phenotype relationships of patients referred to us with suspected arrhythmia syndrome. Methods: Seventeen cases and their twenty relatives were evaluated. Next-generation sequencing analysis was performed for 17 LQTS-related genes. Results: We detected seventeen single nucleotide variants (SNVs) with potential pathogenic significance in 26 of the 36 subjects analyzed. KCNH2 c.172G>A, KCNQ1 c.1768G>A, ANK2 c.4666A>T, c.1484_1485delCT, KCNH2 c.1888G>A were reported as pathogenic or likely pathogenic in HGMD variant classification database. Conclusion: Current study pointed out that early diagnosis can be life-saving for patients and their families by taking family history and detailed examination. Also, we highlight the clinical heterogeneity of arrhythmia syndrome through a patient with a dual phenotype.

Genetic, serological and clinical evaluation of childhood myasthenia syndromes- single center subgroup analysis experience in Turkey.

BACKGROUND: Congenital myasthenic syndrome is a disease that occurs due to several types such as mutations in different pre-synaptic, synaptic, post-synaptic proteins and, glycosylation defects associated with congenital myopathy. Juvenile myasthenia gravis is an autoimmune condition usually caused by antibodies targeting the acetylcholine receptor. AIMS: Our objective is to conduct an analysis on the subgroup traits exhibited by patients who have been diagnosed with congenital myasthenic syndrome and juvenile myasthenia gravis, with a focus on their long-term monitoring and management. METHODS: This study was conducted on children diagnosed with myasthenia gravis, who were under the care of Dokuz Eylul University's Department of Pediatric Neurology for a period of ten years. RESULTS: A total of 22 (12 congenital myasthenic syndrome, 10 juvenile myasthenia gravis) patients were identified. Defects in the acetylcholine receptor (6/12) were the most common type in the congenital myasthenic syndrome group. Basal-lamina-related defects (5/12) were the second most prevalent. One patient had a GFPT1 gene mutation (1/12). Patients with ocular myasthenia gravis (n = 6) exhibited milder symptoms. In the generalized myasthenia gravis group (n = 4), specifically in postpubertal girls, a more severe clinical progression was observed, leading to the implementation of more aggressive treatment strategies. CONCLUSION: This study highlights that clinical recognition of congenital myasthenic syndrome and knowledge of related genes will aid the rapid diagnosis and treatment of these rare neuromuscular disorders. Findings in the juvenile myasthenia gravis group demonstrate the impact of pubertal development and the need for timely and appropriate active therapy, including thymectomy, to improve prognosis.

Genetic evaluation of 50 Turkish patients with neurofibromatosis type 1: 2 years experience of a single center.

BACKGROUND/AIM: Neurofibromatosis type 1 is an autosomal dominant neurocutaneous disorder. Clinical diagnosis is difficult in early childhood, and it is possible to miss a critical interval for tumour screening. In this study, we aimed to characterize the mutational spectrum of Turkish patients and discuss the benefits of molecular testing. MATERIAL AND METHODS: Fifty individuals from 35 unrelated families were included. Main referral reasons for genetic testing were as follows: to confirm a clinical diagnosis, to use in differential diagnosis and to evaluate first-degree family member of a known patient. Two-step process consisting of initial next generation sequencing of the NF1 gene and consequent multiplex ligation-dependent probe amplification were performed. RESULTS: We identified a total of 30 variants in 28 individuals. Variant detection rate was 56% in the entire study group and 71.4% within the index patients. Four novel variants were found. Truncating variants constituted 60% of the entire mutation spectrum. A deletion or duplication was not detected. The most common feature was cafe au lait macules in 70% of the patients, followed by focal areas of signal intensity on brain imaging (26%), cutaneous neurofibromas (24%) and axillary freckling (24%). CONCLUSIONS: Early sequencing in all suspected patients followed by deletion/duplication analysis in patients meeting clinical criteria and a case-to-case based consideration for RNA studies seems to be the effective algorithm for NF-1 diagnosis.

Investigation of different genomic variants in familial Mediterranean fever cases with monoallelic MEFV mutation.

OBJECTIVES: Familial Mediterranean Fever (FMF) is an inflammatory disease characterised by periodic fever and concurrent episodes of serous membrane inflammation. FMF is considered to be inherited in autosomal recessive manner and biallelic mutations in the MEFV gene are associated with the disease. However, approximately 20-25% of patients only have a single mutation in MEFV gene, which creates confusion in differential diagnosis of many patients. This study aimed to reveal rare variants that may act in conjunction with the single pathogenic MEFV variant in the pathogenesis of FMF. METHODS: We performed whole exome sequencing in 17 individuals from 5 different families who were diagnosed according to the clinical criteria, responded positively to colchicine treatment, but had no biallelic MEFV mutation. RESULTS: A disease-causing variant or a common affected cellular pathway that was shared in all index cases was not detected. When cases were examined individually, two de novo variants were identified in the BIRC2 and BCL10 genes, both of which play a role in inflammatory pathways. Functional studies are needed to confirm the physiopathological relationship of these genes with FMF. CONCLUSIONS: This study is one of the most extensive aetiological researches in FMF cases with monoallelic MEFV mutation. We have shown that genotype-phenotype correlation in these cases may not be established by rare genetic variants and discussed underlying causes. Clinical criteria with emphasis on colchicine response and family history should be the main tool and genetic results should only be used for support in FMF diagnosis.

An investigation of the etiology and follow-up findings in 35 children with overgrowth syndromes, including biallelic SUZ12 variant.

Overgrowth-intellectual disability (OGID) syndromes are clinically and genetically heterogeneous group of disorders. The aim of this study was to examine the molecular etiology and long-term follow-up findings of Turkish OGID cohort. Thirty-five children with OGID were included in the study. Single gene sequencing, clinical exome analysis, chromosomal microarray analysis and whole exome sequencing were performed. Five pathogenic copy number variants were detected in the patients; three of them located on chromosome 5q35.2 (encompassing NSD1), others on 9q22.3 and 22q13.31. In 19 of 35 patients; we identified pathogenic variants in OGID genes associated with epigenetic regulation, NSD1 (n = 15), HIST1H1E (n = 1), SETD1B (n = 1), and SUZ12 (n = 2). The pathogenic variants in PIK3CA (n = 2), ABCC9 (n = 1), GPC4 (n = 2), FIBP (n = 1), and TMEM94 (n = 1) which had a role in other growth pathways were detected in seven patients. The diagnostic yield was 31/35(88%). Twelve pathogenic variants were novel. The common facial feature of the patients was prominent forehead. The patients with Sotos syndrome were observed to have milder intellectual disability than patients with other OGID syndromes. In conclusion, this study showed, for the first time, that biallelic variants of SUZ12 caused Imagawa-Matsumoto syndrome, monoallelic variants in SETDIB resulted in OGID. Besides expanded the phenotypes of very rare OGID syndromes caused by FIBP and TMEM94.

Peripheral Expression of ADORA2A Is Increased and Is Correlated with Autism Spectrum Disorder Severity in a Sample of Turkish Children.

Background: The aim of this study was to evaluate the peripheral expression of ADORA2A (Adenosine A2A receptor gene) in young subjects with autism spectrum disorder compared with healthy controls and its relationship with clinical characteristics. Method: This study included 93 children and adolescents with a diagnosis of autism spectrum disorder as the study group and 105 healthy age- and gender-matched controls. Blood samples were obtained from all participants, and a real-time quantitative polymerase chain reaction was performed. Parent- and clinician-rated assessment instruments were used to assess and rate the severity of autism spectrum disorder and other emotional/behavioral problems. Results: The mean age of the study group was 9.06 ± 3.57 and 86% were male (n = 83), whereas the mean age of the control group was 9.22 ± 3.86 and 86.7% were male (n = 91). We have found a higher level of peripheral expression of ADORA2A in children and adolescents with autism spectrum disorder compared with healthy controls (fold change = 1.33, P = .001). We also found a weak negative correlation with autism spectrum disorder severity (r = -0.216; P = .038) and stereotyped behaviors (r = -0.207, P = .046). Conclusion: ADORA2A genes may have a role in the pathophysiology of autism spectrum disorder. Further studies are needed to evaluate whether peripheral expression of ADORA2A genes may be among the biomarkers for diagnosing or measuring the severity of autism spectrum disorder.

Cerebral developmental venous anomalies in children with mismatch repair deficiency.

BACKGROUND: Constitutional mismatch repair deficiency (CMMRD) is one of the rare cancer predisposition syndromes. The aim of this study was to evaluate the cerebral developmental venous anomalies in children with central nervous system tumors associated with CMMRD, an area in which there is extremely little experience. METHODS: Data from children diagnosed with medulloblastoma and high grade central nervous sytem tumor were retrospectively collected. According to the European CMMRD criteria, nine patients were diagnosed as CMMRD syndrome and the others consisted of the group without CMMRD. All radiological examinations of these children were retrospectively reviewed. Whole exome sequencing was performed to index cases` germline DNA. RESULTS: Nine children from four families, six females and three males, were studied. The median age at the first tumor diagnosis was 4.5 years (range, 9 months to 14 years). All CMMRD patients had café au lait spots, but none fulfilled the diagnostic criteria for neurofibromatosis. The patients developed high-grade glial tumor (n: 7) and medulloblastoma (n: 2). The affected genes in the three families were MSH6 [c.478C > T (p.Gln160Ter)], MSH6 [c.2871dupC (p.Phe958LeufsTer5)] and MLH1 [c.236G > A(p.Arg79Lys)], respectively. Seven patients had multiple developmental venous anomalies; six patients had leptomeningeal enhancement; and five patients had cavernomas. None of these findings were present in the group without CMMRD. CONCLUSIONS: Constitutional mismatch repair deficiency should be considered when multiple developmental venous anomalies, cavernomas, and leptomeningeal enhancement are detected, especially in patients with café au lait spots.

Familial clustering of nasopharyngeal carcinoma in the family of an adolescent with nasopharyngeal carcinoma.

BACKGROUND: Nasopharyngeal carcinoma (NPC) is one of the rare malignant diseases of childhood, of which only 1% occurs in children. In recent years, genetic factors have attracted attention in NPC. A very limited data have been reported about clustering within families. CASE: Herein, the familial clustering of nasopharyngeal carcinoma in the family of an adolescent with nasopharyngeal carcinoma is presented. CONCLUSIONS: There is familial clustering in nasopharyngeal carcinoma (NPC), but our knowledge on this subject is limited, especially in children or adolescent populations. Therefore, we should be more careful in NPC in childhood, especially in first-degree relatives.

Hemoglobin A1C can differentiate subjects with GCK mutations among patients suspected to have MODY.

OBJECTIVES: The aim of this study is to determine the clinical and molecular characteristics enabling differential diagnosis in a group of Turkish children clinically diagnosed with MODY and identify the cut-off value of HbA1c, which can distinguish patients with GCK variants from young-onset type 1 and type 2 diabetes. METHODS: The study included 49 patients from 48 unrelated families who were admitted between 2018 and 2020 with a clinical diagnosis of MODY. Clinical and laboratory characteristics of the patients at the time of the diagnosis were obtained from hospital records. Variant analysis of ten MODY genes was performed using targeted next-generation sequencing (NGS) panel and the variants were classified according to American Collage of Medical Genetics and Genomics (ACMG) Standards and Guidelines recommendations. RESULTS: A total of 14 (28%) pathogenic/likely pathogenic variants were detected among 49 patients. 11 variants in GCK and 3 variants in HNF1A genes were found. We identified four novel variants in GCK gene. Using ROC analysis, we found that best cut-off value of HbA1c at the time of diagnosis for predicting the subjects with a GCK variant among patients suspected to have MODY was 6.95% (sensitivity 90%, specificity 86%, AUC 0.89 [95% CI: 0.783-1]). Most of the cases without GCK variant (33/38 [86%]) had an HbA1c value above this cutoff value. We found that among participants suspected of having MODY, family history, HbA1c at the time of diagnosis, and not using insulin therapy were the most differentiating variables of patients with GCK variants. CONCLUSIONS: Family history, HbA1c at the time of diagnosis, and not receiving insulin therapy were found to be the most distinguishing variables of patients with GCK variants among subjects suspected to have MODY.

Congenital Myasthenic Syndromes in Turkey: Clinical and Molecular Characterization of 16 Cases With Three Novel Mutations.

BACKGROUND: Congenital myasthenic syndromes (CMS) are composed of numerous hereditary disorders involving genetic mutations in proteins essential to the integrity of neuromuscular transmission. The symptoms of CMS vary according to the age at onset of symptoms, and the type and severity of muscle weakness. Effective treatment and genetic counseling depend upon the underlying pathogenic molecular mechanism and subtype of CMS. METHODS: A retrospective and cross-sectional study was performed with 16 patients with a genetically confirmed diagnosis of CMS to share our experience with clinical symptoms, demographic data, genetic variants, and treatments applied. RESULTS: Sixteen patients with a specific CMS genetic diagnosis (three novel mutations) were identified, including CHRNE (n = 7), DOK7 (n = 2), AGRN (n = 2), RAPSN (n = 1), CHRNA1 (n = 1), CHRNB1 (n = 1), CHAT (n = 1), and SCN4A (n = 1). Age at onset of symptoms ranged from the neonatal period to 12 years. Genetic diagnosis was confirmed between the ages of three months and 17 years. A significant delay was determined between the onset of symptoms and genetic diagnosis of the disease. CONCLUSIONS: This study highlights the importance of genetic testing in CMS. Due to the rarity of CMS, more cases will be recognized and reported as the use of laboratory and genetic testing accelerates. We hope that our experience will grow and contribute further to the literature as clinical follow-up and treatment increase.

Importance of multigene panel test in patients with consanguineous marriage and family history of breast cancer.

Next-generation sequencing (NGS) technology is used to evaluate hereditary cancer risks of patients worldwide; however, information concerning the germline multigene mutational spectrum among patients with breast cancer (BC) with consanguineous marriage (CM) is limited. Therefore, this prospective study aimed to determine the molecular characteristics of patients with BC who were tested with multigene hereditary cancer predisposition NGS panel and to show the effect of CM on cancer-related genes. Patients with BC with or without CM and family history (FH) of BC treated in our breast center were selected according to The National Comprehensive Cancer Network (NCCN) criteria for hereditary BC. In these patients, the analysis of a panel of 33 genes involved in hereditary cancer predisposition was performed after genetic counseling by using NGS. The pathogenic variant (PV) and the variant of uncertain significance (VUS) were found to be 15.8 and 47.4%, respectively. PVs were identified in 10/33 genes in 34 patients; 38.2% in BRCA1/2 genes; 6, 24, and 14% in other high, moderate and low-risk genes, respectively. The CM rate was 17.7% among the 215 patients with BC. The PV rate was 13.2% in patients with CM and 16.4% in patients without CM (P=0.80). When PV and VUS were evaluated together, the PV+VUS ratio was significantly higher in patients with CM and FH of BC than patients without CM and FH of BC (88.2 vs. 63.3%, P=0.045). Analysis of multigene panel provided 9.76% additional PVs in moderate/low-risk genes. The PV rate was similar in patients with BC with or without CM. A high PV+VUS ratio in patients with CM and FH of BC suggests that genes whose importance are unknown are likely to be pathogenic genes later.

Biallelic BICD2 variant is a novel candidate for Cohen-like syndrome.

Heterozygous mutations in Bicaudal D2 Drosophila homolog 2 (BICD2) gene, encodes a vesicle transport protein involved in dynein-mediated movement along microtubules, are responsible for an exceedingly rare autosomal dominant spinal muscular atrophy type 2A which starts in the childhood and predominantly effects lower extremities. Recently, a more severe form, type 2B, has also been described. Here, we present a patient born to a consanguineous union and who suffered from intellectual disability, speech delay, epilepsy, happy facial expression, truncal obesity with tappering fingers, and joint hypermobility. Whole-exome sequencing analysis revealed a rare, homozygous missense mutation (c.731T>C; p.Leu244Pro) in BICD2 gene. This finding presents the first report in the literature for homozygous BICD2 mutations and its association with a Cohen-Like syndrome. Patients presenting with Cohen-Like phenotypes should be further interrogated for mutations in BICD2.

Clinical and genetic studies of thiamine metabolism dysfunction syndrome-4: case series and review of the literature.

Thiamine metabolism dysfunction syndrome-4 (THMD-4) is an autosomal recessive inherited rare disease (OMIM #613710) characterized by febrile illness associated episodic encephalopathy, leading to transient neurological dysfunction and progressive polyneuropathy. We report three patients from two different families with normal development, episodic encephalopathy, gait disorder, progressive chronic polyneuropathy characterized by motor difficulties, distal weakness, and hoarseness (dysphonia). We identified a homozygous missense c.576G>C, p.(Gln192His) variant in the SLC25A19 gene in both families by whole-exome sequencing. Following genetic diagnosis, thiamine replacement therapy was started, and improvement was observed in all affected patients. We highlight the associated phenotypes of an SCL25A19 mutation leading to clinical features of THMD-4.

Evaluation of hereditary/familial breast cancer patients with multigene targeted next generation sequencing panel and MLPA analysis in Turkey.

Breast cancer, a worldwide leading cause of cancer in women, may occur in familial cases. Germline mutations in BRCA1/2 genes are responsible for 15% of the familial cases. With the power of next generation sequencing (NGS) analysis, it is possible to analyze genes related to hereditary susceptibility to breast cancer and investigate the genetic etiology more thoroughly. In this study, we investigated 30 genes identified frequent pathogenic alleles in Turkish population. The study includes 495 unrelated individuals diagnosed with breast cancer who are selected for genetic testing according to NCCN criteria for hereditary breast cancer. All patients were analyzed by NGS for BRCA1/2 genes. Deletion/duplication investigation by Multiplex ligation-dependent probe amplification (MLPA) and massive sequencing of 30 breast cancer-related genes (Oncorisk Gene Panel) were performed in a stepwise manner. BRCA1/2 variants are the most frequent pathogenic variants which are found in 45 of 495 (9.1%) patients. Four previously unreported, novel, pathogenic variants of BRCA2 gene are identified. In four cases, exonic deletions of BRCA1/2 genes are determined and there is no duplication of these genes. NGS panel investigation involving other moderate-high risk genes contributed genetic diagnosis in an extra 39 out of 419 (9.3%) cases. Our study presents the cost effectiveness of the gene panel approach. We suggest that gene panels should be the first-tier genetic testing for hereditary breast cancer and MLPA analysis of BRCA1/2 genes should be investigated as a complementary method of NGS analysis.

Further delineation of familial polycystic ovary syndrome (PCOS) via whole-exome sequencing: PCOS-related rare FBN3 and FN1 gene variants are identified.

AIM: To identify pathogenic rare coding Mendelian/high-effect size variant(s) by whole-exome sequencing in familial polycystic ovary syndrome (PCOS) patients to elucidate PCOS-related pathways. METHODS: Twenty women and their affected available relatives diagnosed with PCOS according to Rotterdam criteria were recruited. Whole-exome sequencing on germ-line DNA from 31 PCOS probands and their affected relatives was performed. Whole-exome sequencing data were further evaluated by pathway and chemogenomics analyses. In-slico analysis of candidate variants were done by VarCards for functional predictions and VarSite for impact on three-dimensional (3D) structures in the candidate proteins. RESULTS: Two heterozygous rare FBN3 missense variants in three patients, and one FN1 missense variant in one patient from three different PCOS families were identified. CONCLUSION: We identified three novel FBN3 and FN1 variants for the first time in the literature and linked with PCOS. Further functional studies may identify causality of these newly discovered PCOS-related variants, and their role yet remains to be investigated. Our findings may improve our understanding of the biological pathways affected and identify new drug targets.