Whole Exome Sequencing Technique Research Articles

A Comprehensive Overview of NF1 Mutations in Iranian Patients.

Neurofibromatosis type 1 (NF1) is a genetic disorder caused by mutations in the NF1 gene. This disorder shows nearly complete penetrance and high phenotypic variability. We used the whole-exome sequencing technique to identify mutations in 32 NF1 cases from 22 Iranian families. A total of 31 variants, including 30 point mutations and one large deletion, were detected. In eight cases, variants were inherited, while they were sporadic in the remaining. Seven novel variants, including c.5576T > G, c.6658_6659insC, c.2322dupT, c.92_93insAA, c.4360C > T, c.3814C > T, and c.4565_4566delinsC, were identified. The current study is the largest in terms of the sample size of Iranian NF1 cases with identified mutations. The results can broaden the spectrum of NF1 mutations and facilitate the process of genetic counseling in the affected families.

Investigating the Genetic Cause of Ichthyosis Disease in a Family with Exome Sequencing

Background: Ichthyosis is a diverse skin disease characterized by dry skin and itching, particularly affecting the hands. Most forms of this condition are hereditary. Objectives: In this study, we investigated potential new mutations associated with this disease using the new technique of whole-exome sequencing. Methods: In this study, after collecting the patient's blood sample in an EDTA tube and extracting DNA using the salting-out method, next-generation sequencing was performed using the WES method. Following the analysis of the results and identification of the candidate gene, PCR technique and Chromas software were used to confirm the findings. Results: The results indicated the presence of a new variant, NM_001099287: Exon 6:c.C1083A: p.Y361X, in the NIPAL4 gene in a heterozygous form in the parents and a homozygous form in the patient. This variant occurred as a nucleotide substitution in the exon 6 region of the NIPAL4 mRNA sequence. Conclusions: Using the WES technique, it is possible to investigate the presence of new variants related to ichthyosis in a short time and at a low cost.

Complex genotype–phenotype correlation of MYH11: new insights from monozygotic twins with highly variable expressivity and outcomes

BackgroundThoracic aortic aneurysm/dissection (TAAD) and patent ductus arteriosus (PDA) are serious autosomal-dominant diseases affecting the cardiovascular system. They are mainly caused by variants in the MYH11 gene, which encodes the heavy chain of myosin 11. The aim of this study was to evaluate the genotype–phenotype correlation of MYH11 from a distinctive perspective based on a pair of monozygotic twins.MethodsThe detailed phenotypic characteristics of the monozygotic twins from the early fetal stage to the infancy stage were traced and compared with each other and with those of previously documented cases. Whole-exome and Sanger sequencing techniques were used to identify and validate the candidate variants, facilitating the analysis of the genotype–phenotype correlation of MYH11.ResultsThe monozygotic twins were premature and presented with PDA, pulmonary hypoplasia, and pulmonary hypertension. The proband developed heart and brain abnormalities during the fetal stage and died at 18 days after birth, whereas his sibling was discharged after being cured and developed normally post follow-up. A novel variant c.766 A > G p. (Ile256Val) in MYH11 (NM_002474.2) was identified in the monozygotic twins and classified as a likely pathogenic variant according to the American College of Medical Genetics/Association for Molecular Pathology guidelines. Reviewing the reported cases (n = 102) showed that the penetrance of MYH11 was 82.35%, and the most common feature was TAAD (41.18%), followed by PDA (22.55%), compound TAAD and PDA (9.80%), and other vascular abnormalities (8.82%). The constituent ratios of null variants among the cases with TAAD (8.60%), PDA (43.8%), or compound TAAD and PDA (28.6%) were significantly different (P = 0.01). Further pairwise comparison of the ratios among these groups showed that there were significant differences between the TAAD and PDA groups (P = 0.006).ConclusionThis study expands the mutational spectrum of MYH11 and provides new insights into the genotype–phenotype correlation of MYH11 based on the monozygotic twins with variable clinical features and outcomes, indicating that cryptic modifiers and complex mechanisms beside the genetic variants may be involved in the condition.

Expanding the clinical and genetic landscape of (developmental) epileptic encephalopathy with spike-and-wave activation in sleep: results from studies of a Turkish cohort.

The terms developmental epileptic encephalopathy with spike-and-wave activation in sleep (DEE-SWAS) and epileptic encephalopathy with spike-and-wave activation in sleep (EE-SWAS) designate a spectrum of conditions that are typified by different combinations of motor, cognitive, language, and behavioral regression linked to robust spike-and-wave activity during sleep. In this study, we aimed at describing the clinical and molecular findings in "(developmental) epileptic encephalopathy with spike-and-wave activation in sleep" (D)EE-SWAS) patients as well as at contributing to the genetic etiologic spectrum of (D)EE-SWAS. Single nucleotide polymorphism (SNP) array and whole-exome sequencing (WES) techniques were used to determine the underlying genetic etiologies. Of the 24 patients included in the study, 8 (33%) were female and 16 (67%) were male. The median age at onset of the first seizure was 4years and the median age at diagnosis of (D)EE-SWAS was 5years. Of the 24 cases included in the study, 13 were compatible with the clinical diagnosis of DEE-SWAS and 11 were compatible with the clinical diagnosis of EE-SWAS. Abnormal perinatal history was present in four cases (17%), and two cases (8%) had a family history of epilepsy. Approximately two-thirds (63%) of all patients had abnormalities detected on brain computerized tomography/magnetic resonance (CT/MR) imaging. After SNP array and WES analysis, the genetic etiology was revealed in 7 out of 24 (29%) cases. Three of the variants detected were novel (SLC12A5, DLG4, SLC9A6). This study revealed for the first time that Smith-Magenis syndrome, SCN8A-related DEE type 13 and SLC12A5 gene variation are involved in the genetic etiology of (D)EE-SWAS. (D)EE-SWAS is a genetically diverse disorder with underlying copy number variations and single-gene abnormalities. In the current investigation, rare novel variations in genes known to be related to (D)EE-SWAS and not previously reported genes to be related to (D)EE-SWAS were discovered, adding to the molecular genetic spectrum. Molecular etiology enables the patient and family to receive thorough and accurate genetic counseling as well as a personalized medicine approach.

Investigating Mutations of 288 Genes in a Family with Charcot-Marie-Tooth Disease by the Whole-Exome Sequencing Method in Khuzestan Province

Background: Genetic factors play an important role in the occurrence of many diseases, including skeletal diseases. Skeletal diseases are sometimes caused by a mutation in a gene that passes on to children according to the Mendelian inheritance pattern, and sometimes several genes are involved in this path and cause heterogeneous disease. The whole-exome sequencing (WES) technique can be a powerful tool in diagnosing this disease. Methods: Peripheral blood samples (5 mL) were collected from the individuals in EDTA tubes. After sequencing the patient's genome using the WES technique, the candidate genes were checked using the polymerase chain reaction (PCR) technique and Chromas software to confirm the variant in the family. Results: After studying the genome using the WES method and analyzing the results, mutation NM_001005373.4:c.1985C>T was identified by creating a nonsense change in the LRSAM1 gene. Conclusions: The data obtained by the WES technique can help diagnose the disease faster. In addition, these data can be used as background information in the development of a special panel to diagnose this disorder.

Identification of Genetic Variants Associated with Severe Myocardial Bridging through Whole-Exome Sequencing.

Myocardial bridging (MB) is a congenital coronary artery anomaly and an important cause of angina. The genetic basis of MB is currently unknown. This study used a whole-exome sequencing technique and analyzed genotypic differences. Eight coronary angiography-confirmed cases of severe MB and eight age- and sex-matched control patients were investigated. In total, 139 rare variants that are potentially pathogenic for severe MB were identified in 132 genes. Genes with multiple rare variants or co-predicted by ClinVar and CADD/REVEL for severe MB were collected, from which heart-specific genes were selected under the guidance of tissue expression levels. Functional annotation indicated significant genetic associations with abnormal skeletal muscle mass, cardiomyopathies, and transmembrane ion channels. Candidate genes were reviewed regarding the functions and locations of each individual gene product. Among the gene candidates for severe MB, rare variants in DMD, SGCA, and TTN were determined to be the most crucial. The results suggest that altered anchoring proteins on the cell membrane and intracellular sarcomere unit of cardiomyocytes play a role in the development of the missed trajectory of coronary vessels. Additional studies are required to support the diagnostic application of cardiac sarcoglycan and dystroglycan complexes in patients with severe MB.

A novel causative functional mutation in GATA6 gene is responsible for familial dilated cardiomyopathy as supported by in silico functional analysis

Dilated cardiomyopathy (DCM), one of the most common types of cardiomyopathies has a heterogeneous nature and can be seen in Mendelian forms. Next Generation Sequencing is a powerful tool for identifying novel variants in monogenic disorders. We used whole-exome sequencing (WES) and Sanger sequencing techniques to identify the causative mutation of DCM in an Iranian pedigree. We found a novel variant in the GATA6 gene, leading to substituting Histidine by Tyrosine at position 329, observed in all affected family members in the pedigree, whereas it was not established in any of the unaffected ones. We hypothesized that the H329Y mutation may be causative for the familial pattern of DCM in this family. The predicted models of GATA6 and H329Y showed the high quality according to PROCHECK and ERRAT. Nonetheless, simulation results revealed that the protein stability decreased after mutation, while the flexibility may have been increased. Hence, the mutation led to the increased compactness of GATA6. Overall, these data indicated that the mutation could affect the protein structure, which may be related to the functional impairment of GATA6 upon H329Y mutation, likewise their involvement in pathologies. Further functional investigations would help elucidating the exact mechanism.

Familial exudative vitreoretinopathy in a 4 generations family of South-East Asian Descendent with FZD4 mutation (c.1501_1502del)

BackgroundFamilial Exudative Vitreoretinopathy (FEVR) is a hereditary disorder characterized by peripheral avascular retina with neovascularization. Although FEVR has been thoroughly described in multiple literature publications from different countries, there are currently limited articles describing the phenotypes of FEVR among South-East Asian Descendent. This paper describes the clinical phenotype of the FZD4 gene with c.1501_1502 deletion in a 4-generation case series of a South East Asian family.MethodsWe reviewed a 4-generation case series of a South-East Asian descendent family consisting of 27 family members with 10 members diagnosed with FEVR. We observed the clinical phenotype of these series of patients, including some of the family members who underwent whole-exome sequencing, PCR amplification and DNA sequencing techniques to identify the mutated gene.ResultsFrameshift mutation (c.1501_1502del) were found in FZD4 gene in this series of patients with the age ranging from 1 month old to 69 years old. There was a 100% (4/4) of our paediatric patients being diagnosed within 21 days of life. It was also found that 75% of patients (6/8) less than 40 years old exhibited disease asymmetry of 2 stages or more and 80% (8/10) had a history of vitreoretinal surgery or diode laser photocoagulation, with a further 50% of the adult patients identified as legally blind; the mean age of blindness was 18-years-old.ConclusionsPhenotypic manifestation of FZD4 gene with c.1501_1502del mutation can be identified within the neonatal period. They have relatively greater clinical asymmetry of 2 stages or more compared to other mutations. Without treatment, most of them will have bilateral severe visual impairment around the adolescent age group.

MO1057: Cost-Analysis of a Clinical Workflow for Diagnosis of Inherited Kidney Diseases

Abstract BACKGROUND AND AIMS In the last decade, the use of whole-exome sequencing techniques (WES) has provided many insights into inherited kidney diseases that are thought to represent at least 10%–15% of cases of end-stage CKD [1]. However, among others, cost concerns limit the widespread of genomics use in daily practice [1, 2]. Publicly funded genomic testing is restricted in most health care systems [2]. Consequently, the evaluation of cost-effectiveness is urgently required in order to establish genomic sequencing as a standard diagnostic test for nephropathic patients. The aim of this study was to perform a cost-analysis of genetic testing use in a diagnostic workflow. METHOD We recently set up a diagnostic workflow for the selection of patients that should undergo genetic testing in the suspicion of a genetic disease. This algorithm is applied by a network of nephrology centres on the regional territory. Selected patients are referred to a tertiary centre, Meyer University Hospital of Florence (Italy), for genetic diagnosis by WES. We enrolled paediatric and adult patients referred to the outpatient service based on the pre-specified clinical criteria. All the patients underwent genetic testing from 2018 to June 2021. We conducted a cost-analysis in two parts: (1) assessment of the cost-effectiveness cut-off; and (2) exploratory modeled cost-analysis using WES in different phases of the diagnostic trajectory. As a surrogate of the cost-effectiveness analysis, we calculated the cost-effectiveness cut-off, indicating the amount of expenses for diagnostic examinations at which WES sequencing would be cost-effective. For the exploratory cost-analysis, we defined two diagnostic trajectories: Model I, considering an ideal complete diagnostic pathway and late use of WES; and Model II, considering an early use of WES allowing to save a certain number of examinations. Genomic and non-genomic investigations were obtained from local practice, available clinical evidence and guidelines. We then calculated the cost per diagnosis according to each Model for different clinical categories and for the whole study population using the diagnostic rates of this study. We considered only direct medical costs, based on the regional health reimbursement system, which is comprehensive of materials and human resources. The prices are expressed in euros. The analysis was conducted from a regional healthcare system perspective. RESULTS The analysis included 402 patients. WES performed after the standard non-conclusive diagnostic work-up (at a mean cost of €2992/patient) resulted in cost-effectiveness at a cost of <€2004/patient in the study population. Looking at different clinical categories, WES was cost-effective at a cost ranging from €3336 to €770. Overall, the exploratory cost-analysis showed Model II as cost reducing in comparison to Model I. The mean cost per diagnosis in Model I with late use of WES was estimated at €7700. Early use of WES in Model II had an estimated cost per diagnosis of €6340, leading to a cost saving of €1360/patient tested. Regarding the single clinical categories, the highest cost saving per diagnosis was obtained in podocytopathies (Model I: €13.010 versus Model II: €7261). The early use of WES produced a slight increase in estimated costs per diagnosis for tubulopathies, as well as for ciliopathies, while for syndromic CKD and metabolic kidney disorders, the costs per diagnosis did not result in major changes. CONCLUSION Early use of WES in the diagnostic pathway of inherited diseases, guided by a framework of specific criteria, is feasible and has the potential to produce substantial cost savings in healthcare.

FC036: A Clinical Workflow for Selection of Patients and Cost-Efficient Diagnosis of Genetic Kidney Diseases

Abstract BACKGROUND AND AIMS With the spread of whole-exome sequencing techniques (WES), genetic kidney diseases are increasingly recognized across all age groups. However, accessibility, interpretation of results and costs limit the widespread of genomics use in daily practice [1]. In the very last years, first experiences in the implementation of renal genetic services have been started [2]. In this work, we explored the feasibility and diagnostic performance of a service delivery model based on a territorial network for patients’ selection, followed by referral to the renal genetic clinic of a tertiary centre for WES, results interpretation and counseling. We hypothesized that higher diagnostic yield and cost-effectiveness could be achieved by implementing a workflow that could solve three problems: (1) Give clear indications to the nephrologist about which patients with kidney diseases should undergo genetic testing, (2) Increase the rate and accuracy of genetic testing and evaluate the clinical impact and (3) Identify the kidney diseases for which early genetic testing is not only clinically, but also economically convenient for the healthcare provider based on pre-specified clinical criteria for tertiary centre referral. METHOD We set up a multi-step diagnostic workflow. We established specific clinical criteria for evaluation of patients who should undergo genetic testing on the suspicion of a genetic disease by a network of nephrology centres. Patients selected were referred to a tertiary centre for genetic diagnosis by WES, reverse-phenotyping and multidisciplinary board analysis. This workflow has been applied to pediatric and adult patients with kidney diseases belonging to eight clinical categories (Podocytopathies, Collagenopathies, Tubulopathies, Unknown familial nephropathies, Ciliopathies, Congenital anomalies of the kidney and urinary tract, Syndromic chronic kidney disease and Metabolic kidney disorders). We recorded clinical-laboratory-radiological information of patients included. We also performed a cost-analysis of the diagnostic workflow modelling the possible economic saving using it. RESULTS We included 402 patients, of note, 188 patients were female (46.8%) and 132 patients (33%) were >16 years of age. We obtained a global diagnostic yield of 69.2% (278–402), with category-specific diagnostic rates ranging from 38.5% to 87%. By reverse phenotyping, we reclassified diagnoses in 74–278 (26.6%) patients, thus increasing diagnostic accuracy. Overall, reverse phenotyping increased the diagnostic rate in an average of 20% of cases, irrespective of the age at clinical onset of the disease. Diagnostic yield was independent of the age at the onset of kidney disease. Genetic testing was offered as cascade screening to 67 families, providing a genetic diagnosis in 62 family members with previously unsuspected or unspecified kidney disorders. The clinical work-up changed and was redirected on average in 50% of patients. In 11.5% of patients, the results of genetic testing helped in guiding kidney transplant decisions. Finally, cost-analysis showed that our workflow is cost-efficient allowing to potentially save a mean of 1360 euros per patient. CONCLUSION Ordering genetic testing, interpreting results, counselling patients and their families, and tailoring clinical management (i.e. personalized nephrology) is feasible and saves costs in a real-world setting.

Whole exome sequencing is an alternative method in the diagnosis of mitochondrial DNA diseases.

BackgroundMitochondrial disease (MD) is genetically a heterogeneous group of disorders with impairment in respiratory chain complexes or pathways associated with the mitochondrial function. Nowadays, it is still a challenge for the genetic screening of MD due to heteroplasmy of mitochondrial genome and the complex model of inheritance. This study was designed to investigate the feasibility of whole exome sequencing (WES)‐based testing as an alternative option for the diagnosis of MD.MethodsA Chinese Han cohort of 48 patients with suspect MD features was tested using nanoWES, which was a self‐designed WES technique that covered the complete mtDNA genome and 21,019 nuclear genes. Fourteen patients were identified with a single genetic variant and three with single deletion in mtDNA.ResultsThe heteroplasmy levels of variants in mitochondrial genome range from 11% to 100%. NanoWES failed to identify multiple deletions in mtDNA compared with long range PCR and massively parallel sequencing (LR‐PCR/MPS). However, our testing showed obvious advantages in identifying variations in nuclear DNA. Based on nanoWES, we identified two patients with nuclear DNA variation. One of them showed Xp22.33‐q28 duplication, which indicated a possibility of Klinefelter syndrome.ConclusionNanoWES yielded a diagnostic rate of 35.4% for MD. With the rapid advances of next generation sequencing technique and decrease in cost, we recommend the usage of nanoWES as a first‐line method in clinical diagnosis.

Pedigree Analysis of ACTN1-Related Thrombocytopenia Attributed to A Novel Mutation

All the family members' peripheral blood was collected for routine blood tests, blood smear, coagulation function, and platelet aggregation test. Flow cytometry was used to detect the expression of platelet CD41 and CD61. The proband and her father were tested bone marrow cytomorphology. Whole-exome sequencing techniques were performed to detect and uncover mutant loci of suspected pathogenic genes. Bioinformatics was used to assess the conserved nature of the mutated loci and to analyze the effect of the mutated genes leading to the function of the corresponding amino acid sequences. The platelet count of the proband was 88×109/L, and the blood smear showed dumbbell-shaped platelets, snake-shaped platelets and platelets of various sizes. Her bone marrow cytomorphology revealed normal megakaryocyte morphology with a count of 270. The platelet count of the proband's father was 74×109/L, with large platelets and platelets of various sizes observed in the blood smear, and the morphology of megakaryocytes was normal in bone marrow with a megakaryocyte count of 239. Her grandfather had a platelet count of 83×109/L, with snake-shaped platelets and platelets of various sizes on blood smears. Other family members were normal in all tests. The missense mutation c.2396G > A in exon 20 of the ACTN1 gene in the proband resulted in the mutation of 799 amino acids of the encoded protein, i.e., Arg, to His. The sequencing results of her father and grandfather at this locus were found to be consistent with her. Furthermore, bioinformatics analysis indicated that the locus was highly conserved across species and that variation in this locus might lead to functional impairment of the protein. The protein model analysis demonstrated that α-actin-1 at position 799 Arg and Glu at position 811 could form a critical salt bridge which stabilizes the conformation of the Ca2+ binding loop within the calmodulin-like motif. the mutation of R799H lost this critical salt bridge and destabilized this structural domain. In the present study, the newly uncovered missense mutation c.2396G＞A in exon 20 of the ACTN1 gene is potentially the molecular mechanism for the thrombocytopenia.

Clinical, radiological, and molecular diagnosis of congenital pituitary diseases causing short stature

Short stature in children can be caused by congenital pituitary disorders involving at least one form of growth hormone deficiency. Clinical and radiological evaluations of the index case and family history assessments are essential to guide genetic diagnostic testing and interpret results. The first-line approach is panel testing of genes involved in pituitary development with variants known to be pathogenic in this context. It identifies a genetic cause in less than 10% of cases, however. Whole-exome and whole-genome sequencing techniques may provide original information but also raise new questions regarding the pathophysiological role of identified variants. These new tools can make genetic counselling more complex. The role of clinicians in these interpretations is therefore important. © 2022 French Society of Pediatrics. Published by Elsevier Masson SAS. All rights reserved.

Common and Unique Genetic Background between Attention-Deficit/Hyperactivity Disorder and Excessive Body Weight.

Comorbidity studies show that children with ADHD have a higher risk of being overweight and obese than healthy children. This study aimed to assess the genetic alternations that differ between and are shared by ADHD and excessive body weight (EBW). The sample consisted of 743 Polish children aged between 6 and 17 years. We analyzed a unique set of genes and polymorphisms selected for ADHD and/or obesity based on gene prioritization tools. Polymorphisms in the KCNIP1, SLC1A3, MTHFR, ADRA2A, and SLC6A2 genes proved to be associated with the risk of ADHD in the studied population. The COMT gene polymorphism was one that specifically increased the risk of EBW in the ADHD group. Using the whole-exome sequencing technique, we have shown that the ADHD group contains rare and protein-truncating variants in the FBXL17, DBH, MTHFR, PCDH7, RSPH3, SPTBN1, and TNRC6C genes. In turn, variants in the ADRA2A, DYNC1H1, MAP1A, SEMA6D, and ZNF536 genes were specific for ADHD with EBW. In this way, we confirmed, at the molecular level, the existence of genes specifically predisposing to EBW in ADHD patients, which are associated with the biological pathways involved in the regulation of the reward system, intestinal microbiome, and muscle metabolism.

Waardenburg syndrome type II in a Chinese pedigree caused by frameshift mutation in the SOX10 gene.

Waardenburg syndrome (WS) is a congenital hereditary disease, attributed to the most common symptoms of sensorineural deafness and iris hypopigmentation. It is also known as the hearing-pigmentation deficient syndrome. Mutations on SOXl0 gene often lead to congenital deafness and has been shown to play an important role in the pathogenesis of WS. We investigated one family of five members, with four patients exhibiting the classic form of WS2, whose DNA samples were analyzed by the technique of Whole-exome sequencing (WES). From analysis of WES data, we found that both the mother and all three children in the family have a heterozygous mutation on the Sex Determining Region Y - Box 10 (SOX10) gene. The mutation was c.298_300delinsGG in exon 2 of SOX10 (NM_006941), which leads to a frameshift of nine nucleotides, hence the amino acids (p. S100Rfs*9) are altered and the protein translation may be terminated prematurely. Further flow cytometry confirmed significant down-regulation of SOX10 protein, which indicated the SOX10 gene mutation was responsible for the pathogenesis of WS2 patients. In addition, we speculated that some other mutated genes might be related to disease phenotype in this family, which might also participate in promoting the progression of WS2.

A Clinical-Based Diagnostic Approach to Cerebellar Atrophy in Children

Background: Cerebellar atrophy is a neuroradiological definition that categorizes conditions heterogeneous for clinical findings, disease course, and genetic defect. Most of the papers proposing a diagnostic workup for pediatric ataxias are based on neuroradiology or on the literature and experimental knowledge, with a poor participation of clinics in the process of disease definition. Our study aims to offer a different perspective on the way we approach cerebellar atrophy in developmental age, building a clinical-based diagnostic workup to guide molecular diagnosis. Methods: we recruited 52 patients with pediatric-onset cerebellar atrophy and definite disease categorization. Children underwent brain MRI, neurophysiological exams, metabolic investigations, and muscle biopsy with respiratory chain complex study. Single-gene sequencing, next-generation sequencing NGS panels, whole-exome sequencing (WES), and disease-specific techniques have been used to reach genetic confirmation. Results: Brain MRI is the main method of diagnosis, followed by tests on muscle biopsy and peripheral nervous system study. Other exams (e.g., metabolic investigations or evoked potentials) may be useful to narrow the list of diagnostic possibilities. Conclusions: We propose a diagnostic approach to cerebellar atrophy in children based on clinical findings, and support the evidence that a precise phenotypic definition may lead to the formulation of a definite diagnosis or otherwise guide the back phenotyping process derived from large molecular data.

Updates in the advances of sporadic medullary thyroid carcinoma: from the molecules to the clinic.

Medullary thyroid carcinoma (MTC) is a rare neuroendocrine malignancy that originates in parafollicular cells. It is well-known that a quarter of MTC are involved in hereditary multiple endocrine neoplasia type 2 syndromes, whereas most MTC are sporadic. Unlike the commonly encountered gastrointestinal or pulmonary neuroendocrine tumors, most sporadic MTCs have distinct genetic alterations featured by somatic changes of either Rearranged during Transfection (RET) or RAS point mutation. The increasing application of next-generation sequencing, whole-exome sequencing, and other molecular detection techniques enables us to understand MTC comprehensively concerning its detailed molecular changes and their clinical correlations. This article reviews the advances in genetic alterations and their prognostic impact in sporadic MTC among different populations and discusses the associated tumor immune microenvironments and the potential role of immunotherapy targeting PD-L1/PD-1 in treating MTC. Furthermore, the current multikinase inhibitor targeting therapy for sporadic MTC has been summarized here and its efficacy and drug toxicity are discussed. Updates in advance of the role of calcitonin/procalcitonin/calcitonin-related polypeptide alpha (CALCA) gene transcripts in diagnosing and handling MTC are also mentioned. The treatment of advanced MTC is still challenging and might require a combination of several modalities.

A novel missense mutation (c.1006C>T) of SPG20 gene associated with Troyer syndrome

The number of gene mutations involved in the hereditary spastic paraplegias is rapidly growing due to the expansion of the frontiers of genomic research by next-generation DNA sequencing platforms. Nevertheless, a comprehensive genetic diagnosis method remains yet unavailable for these diseases. In the current research, an 8-year-old boy with short stature and developmental delay impairment, from a nonconsanguineous family, was referred to our genetic lab. Firstly, based on the physician recommendation, the patient was evaluated by tandem mass spectrometry (MS/MS) for the quantitative examination of amino acids, and then the patient was genetically investigated by karyotype analysis and whole-exome sequencing (WES) technique. Subsequently, targeted Sanger sequencing was applied to confirm the presence of the candidate variant in all the members of the family and screening the other patients for Troyer syndrome. Analysis of inherited metabolic disorders by tandem MS/MS showed the state of all the family members as normal and also karyotyping indicated no chromosomal aberration in the patient. Further investigation by WES technique indicated a homozygous missense variant in the SPG20 gene, c.1006C[T. Targeted sequencing result of the mutation confirmed homozygote state for the affected case and a heterozygote genotype for his parents. The mutation was classified as pathogenic. Detection of novel variants especially pathogenic variantin the SPG20 gene was associated with Troyer syndrome, which encodes a multifunctional protein termed Spartin, assist in improving genotype-phenotype correlation of genetic variants and may facilitate initial diagnosis of Troyer syndrome.

Emerging therapies in genodermatoses

The human genome project yielded a compendium of genetic material that has allowed rapid advancement both in the technique of whole exome sequencing and also in the ability to identify single gene defects. The next generation of genetics has investigated how these genes interact in the development of disease, identifying pathways of illness and end organ tissue abnormal development. From the knowledge of single genes and pathways of genodermatosis development arises the opportunity to produce genetic therapies. This contribution reviews some of the exciting, emerging genetic therapies in genodermatoses.

Identification of second primary tumors from lung metastases in patients with esophageal squamous cell carcinoma using whole-exome sequencing.

Esophageal squamous cell carcinoma (ESCC) patients with a synchronous or metachronous lung tumor can be diagnosed with lung metastasis (LM) or a second primary tumor (SPT), but the accurate discrimination between LM and SPT remains a clinical dilemma. This study aimed to investigate the feasibility of using the whole-exome sequencing (WES) technique to distinguish SPT from LM.Methods: We performed WES on 40 tumors from 14 patients, including 12 patients with double squamous cell carcinomas (SCCs) of the esophagus and lung (lymph node metastases were sequenced as internal controls) diagnosed as LM according to pathological information and 2 patients with paired primary ESCC and non-lung metastases examined as external controls.Results: Shared genomic profiles between esophageal (T) and lung (D) tumors were observed in 7 patients, suggesting their clonal relatedness, thus indicating that the lung tumors of these patients should be LM. However, distinct genomic profiles between T and D tumors were observed in the other 5 patients, suggesting the possibility of SPTs that were likely formed through independent multifocal oncogenesis.Conclusions: Our data demonstrate the limitations and insufficiency of clinicopathological criteria and that WES could be useful in understanding the clonal relationships of multiple SCCs.