A compound heterozygous mutation of ERCC8 is responsible for a family with Cockayne syndrome.

BACKGROUND: Cockayne syndrome is an inherited heterogeneous defect in transcription-coupled DNA repair (TCR) cause severe clinical syndromes, which may affect the nervous system development of infants and even lead to premature death in some cases. ERCC8 diverse critical roles in the nucleotide excision repair (NER) complex, which is one of the disease-causing genes of Cockayne syndrome. METHODS AND RESULTS: The mutation of ERCC8 in the patient was identified and validated using WES and Sanger sequencing. Specifically, a compound heterozygous mutation (c.454_460dupGTCTCCA p. T154Sfs*13 and c.755_759delGTTTT p.C252Yfs*3) of ERCC8 (CSA) was found, which could potentially be the genetic cause of Cockayne syndrome in the proband. CONCLUSION: In this study, we identified a novel heterozygous mutation of ERCC8 in a Chinese family with Cockayne syndrome, which enlarging the genetic spectrum of the disease.

Next-generation sequencing through multi-gene panel testing for the diagnosis of a Chinese patient with atypical Cockayne syndrome.

BACKGROUND: Cockayne syndrome (CS, OMIM #133540, #216400) is a rare autosomal recessive disease involving multiple systems, typically characterized by microcephaly, premature aging, growth retardation, neurosensory abnormalities, and photosensitivity. The age of onset is related to the severity of the clinical phenotype, which may lead to fatal outcomes. METHODS: We report a 3-year-old girl who presented with photosensitivity, gait abnormalities, stunting, and microcephaly and showed atypical clinical classification due to mild clinical manifestations at an early onset age. RESULTS: Next-generation sequencing reveals the frameshift mutation (c.394_398del, p.Leu132Asnfs*6) and a novel microdeletion of ERCC8 (exon4del, p.Arg92fs). CONCLUSION: Therefore, it is still necessary to carry out next-generation sequencing for CS patients with atypical clinical manifestations, which is essential for diagnosis and accurate genetic counseling.

A Novel Missense Mutation in ERCC8 Co-Segregates with Cerebellar Ataxia in a Consanguineous Pakistani Family.

Autosomal-recessive cerebellar ataxias (ARCAs) are heterogeneous rare disorders mainly affecting the cerebellum and manifest as movement disorders in children and young adults. To date, ARCA causing mutations have been identified in nearly 100 genes; however, they account for less than 50% of all cases. We studied a multiplex, consanguineous Pakistani family presenting with a slowly progressive gait ataxia, body imbalance, and dysarthria. Cerebellar atrophy was identified by magnetic resonance imaging of brain. Using whole exome sequencing, a novel homozygous missense mutation ERCC8:c.176T>C (p.M59T) was identified that co-segregated with the disease. Previous studies have identified homozygous mutations in ERCC8 as causal for Cockayne Syndrome type A (CSA), a UV light-sensitive syndrome, and several ARCAs. ERCC8 plays critical roles in the nucleotide excision repair complex. The p.M59T, a substitution mutation, is located in a highly conserved WD1 beta-transducin repeat motif. In silico modeling showed that the structure of this protein is significantly affected by the p.M59T mutation, likely impairing complex formation and protein-protein interactions. In cultured cells, the p.M59T mutation significantly lowered protein stability compared to wildtype ERCC8 protein. These findings expand the role of ERCC8 mutations in ARCAs and indicate that ERCC8-related mutations should be considered in the differential diagnosis of ARCAs.

Cockayne syndrome without UV-sensitivity in Vietnamese siblings with novel ERCC8 variants.

Cockayne syndrome (CS) is a rare progeroid disorder characterized by growth failure, microcephaly, photosensitivity, and premature aging, mainly arising from biallelic ERCC8 (CS-A) or ERCC6 (CS-B) variants. In this study we describe siblings suffering from classical Cockayne syndrome but without photosensitivity, which delayed a clinical diagnosis for 16 years. By whole-exome sequencing we identified the two novel compound heterozygous ERCC8 variants c.370_371del (p.L124Efs*15) and c.484G>C (p.G162R). The causality of the ERCC8 variants, of which one results in a frameshift and the other affects the WD3 domain, was tested and confirmed by a rescue experiment investigating DNA repair in H2O2 treated patient fibroblasts. Structural modeling of the p.G162R variant indicates effects on protein-protein interaction. This case shows the importance to test for ERCC6 and ERCC8 variants even if patients do not present with a complete CS phenotype.

Whole exome sequencing identifies a novel variant causing cockayne syndrome type I in a consanguineous Pakistani family.

BACKGROUND: Cockayne syndrome (CS) is a rare neurodegenerative disorder characterized by impaired neurological functions, cachectic dwarfism, microcephaly and photosensitivity. Complementation assays identify two groups of this disorder, CS type I (CSA) and CS type II (CSB), caused by mutations in ERCC8 and ERCC6, respectively. OBJECTIVES: This study aimed to investigate the genetic basis of a consanguineous Pakistani family with three affected individuals presenting with typical clinical symptoms of CS. METHODS: We employed whole exome sequencing of the proband and then Sanger sequenced all the family members to confirm its segregation in the family. Different bioinformatics tools were used to predict pathogenicity of this variant. RESULTS: Variants were filtered according to the pedigree structure. We identified a novel homozygous variant (c.202A>T; p.Ile68Phe) in ERCC8 gene in the proband. The variant was found to segregate in the family. CONCLUSIONS: These findings add to the genetic heterogeneity of ERCC8 and expands the mutation spectrum. Also, identification of this variant can facilitate prenatal diagnosis/genetic counselling set ups in Pakistan where this disease largely remains undiagnosed.

Heterogeneous clinical features in Cockayne syndrome patients and siblings carrying the same CSA mutations.

BACKGROUND: Cockayne syndrome (CS) is a rare autosomal recessive disorder caused by mutations in ERCC6/CSB or ERCC8/CSA that participate in the transcription-coupled nucleotide excision repair (TC-NER) of UV-induced DNA damage. CS patients display a large heterogeneity of clinical symptoms and severities, the reason of which is not fully understood, and that cannot be anticipated in the diagnostic phase. In addition, little data is available for affected siblings, and this disease is largely undiagnosed in North Africa. METHODS: We report here the clinical description as well as genetic and functional characterization of eight Tunisian CS patients, including siblings. These patients, who belonged to six unrelated families, underwent complete clinical examination and biochemical analyses. Sanger sequencing was performed for the recurrent mutation in five families, and targeted gene sequencing was done for one patient of the sixth family. We also performed Recovery RNA Synthesis (RRS) to confirm the functional impairment of DNA repair in patient-derived fibroblasts. RESULTS: Six out of eight patients carried a homozygous indel mutation (c.598_600delinsAA) in exon 7 of ERCC8, and displayed a variable clinical spectrum including between siblings sharing the same mutation. The other two patients were siblings who carried a homozygous splice-site variant in ERCC8 (c.843+1G>C). This last pair presented more severe clinical manifestations, which are rarely associated with CSA mutations, leading to gastrostomy and hepatic damage. Impaired TC-NER was confirmed by RRS in six tested patients. CONCLUSIONS: This study provides the first deep characterization of case series of CS patients carrying CSA mutations in North Africa. These mutations have been described only in this region and in the Middle-East. We also provide the largest characterization of multiple unrelated patients, as well as siblings, carrying the same mutation, providing a framework for dissecting elusive genotype-phenotype correlations in CS.

Identification of ERCC8 as a novel cisplatin-resistant gene in esophageal cancer based on genome-scale CRISPR/Cas9 screening.

Esophageal cancer (ESCA) is one of the most common malignant tumors of the digestive system worldwide. As a first-line drug for chemotherapy, cisplatin resistance is the major obstacle in the successful treatment of esophageal cancer. Previous studies largely failed to identify the key genes associated with cisplatin resistance. Hence, the aim of this study was to screen the cisplatin resistance-related genes of esophageal cancer using CRISPR/Cas9 gene-editing technology and Brunello iBar library. Of note, we identified ERCC8 as a novel cisplatin-resistant gene by high-throughput sequencing and cisplatin resistance assays. Based on KEGG and GO analysis, we hypothesized that the mechanism of ERCC8 involvement in cisplatin resistance is through binding to damaged DNA to perform nucleotide excision repair, contributing to the restoration of basic DNA functions and cellular life activities in ESCA. In addition, Cell proliferation and wound healing assay confirmed that ERCC8 had little effect on the proliferation and migration of esophageal cancer cells in vitro. Survival analysis showed that ERCC8 expression was not associated with OS, DSS, or FPI in patients with ESCA. Immuno-infiltration analysis indicated that increased ERCC8 expression is associated with NK cells, macrophages, T helper cells, Th1 cells, and Th2 cells. Collectively, ERCC8 may serve as a new biomarker for predicting cisplatin resistance and have the prospect of becoming an effective target for the clinical treatment of cisplatin resistance in ESCA.

Identification of two novel homozygous mutations in ERCC8 gene in two unrelated consanguineous families with Cockayne syndrome from Iran.

BACKGROUND: Cockayne syndrome (CS) is a rare autosomal recessive disorder with characteristic multisystem involvement including pre- or post-natal growth failure, progressive neurological dysfunction, psychomotor retardation, cerebral atrophy, microcephaly and mental retardation, due to mutations in either the ERCC8/CSA or ERCC6/CSB gene. METHOD: We present two Iranian patients with remarkable growth failure, developmental delay, microcephaly, severe speech delay, vision problem, sun sensitivity, hearing loss, dental anomalies, unstable gait, mild contractures in knees, kyphosis and spasticity in lower limbs, balance disorders and typical dysmorphic features including long nose, aged face, large ears and sunken eyes. Clinical evaluation, magnetic resonance imaging, Peripheral blood karyotype, Multiplex ligation-dependent probe amplification (MLPA), and whole-exome sequencing were used to characterize etiology in two patients from two unrelated consanguineous families of Iranian descent with Cockayne syndrome. RESULTS: We detected two novel pathogenic mutations in two unrelated families, a homozygous duplication mutation (c.317_320dupAGTG, p.Trp107Ter) and a splicing variant (c.481 + 1G > A) in ERCC8 gene. CONCLUSION: WES results together with the characteristic clinical manifestations of Cockayne syndrome, provided an accurate diagnosis for two patients. Also, our study identified two novel variants in Iranian families.

Associations of individual and joint expressions of ERCC6 and ERCC8 with clinicopathological parameters and prognosis of gastric cancer.

BACKGROUND: Excision repair cross-complementing group 6 and 8 (ERCC6 and ERCC8) have been implicated in ailments such as genetic diseases and cancers. However, the relationship between individual and joint expressions of ERCC6/ERCC8 and clinicopathological parameters as well as prognosis of gastric cancer (GC) still remains unclear. METHODS: In this study, protein expressions of ERCC6, ERCC8 and ERCC6-ERCC8 were detected by immunohistochemistry (IHC) in 109 paired GC and para-cancerous normal tissue samples. The mRNA expression was detected in 36 pairs of tissue samples. IHC results and RNA-seq data extracted from The Cancer Genome Atlas (TCGA) were used to explore the clinical value of ERCC6 and ERCC8 expression in GC. We further conducted protein-protein interaction analysis, Gene Ontology, Kyoto Encyclopedia of Genes and Genomes, gene set enrichment analysis, and gene-gene interaction analysis to explore the function and regulation networks of ERCC6 and ERCC8 in GC. RESULTS: Individual and joint ERCC6/ERCC8 expression were significantly higher in adjacent normal mucosa compared with GC tissues. ERCC6 mRNA expression showed no difference in GC and paired tissues, while ERCC8 mRNA was significantly decreased in GC tissues. Protein expression of ERCC6, ERCC8, double negative ERCC6-ERCC8 and double positive ERCC6-ERCC8 and overexpressed ERCC6 mRNA were related to better clinicopathologic parameters, while overexpressed ERCC8 mRNA suggested worse parameters. Univariate survival analysis indicated that the OS was longer when ERCC6 protein expression and ERCC8 mRNA expression increased, and double negative ERCC6-ERCC8 expression was associated with a short OS. Bioinformatics analyses showed ERCC6 and ERCC8 were associated with nucleotide excision repair (NER) pathway, and six and ten gene sets were figured out to be related with ERCC6 and ERCC8, respectively. KEGG pathway showed that ERCC6/ERCC8 related gene sets were mainly involved in the regulation of PI3K/AKT/mTOR pathway. Direct physical interactions were found between ERCC6 and ERCC8. CONCLUSIONS: Individual and joint expressions of ERCC6/ERCC8 were associated with clinical features of GC. Protein expression of ERCC6, ERCC6-ERCC8, and mRNA expression of ERCC8 were related to prognosis of GC. ERCC6 and ERCC8 primarily function in the NER pathway, and may regulate GC progression through the regulation of PI3K/AKT/mTOR pathway.

Clinical and Mutation Spectra of Cockayne Syndrome in India.

BACKGROUND: Cockayne syndrome is an autosomal recessive disorder caused by biallelic mutations in ERCC6 or ERCC8 genes. AIMS: To study the clinical and mutation spectrum of Cockayne syndrome. SETTING AND DESIGN: Medical Genetics Outpatient Department of Sanjay Gandhi Post Graduate Institute of Medical Sciences, Lucknow. This was a prospective study from 2007 to 2015. MATERIALS AND METHODS: Clinical details were recorded, and sequencing of ERCC6 and ERCC8 were performed. RESULTS AND CONCLUSIONS: Of the six families, one family had a homozygous mutation in ERCC8 and the other five families had homozygous mutations in ERCC6. Novel variants in ERCC6 were identified in four families. Phenotypic features may vary from severe to mild, and a strong clinical suspicion is needed for diagnosis during infancy or early childhood. Hence, molecular diagnosis is needed for confirmation of diagnosis in a child with a suspicion of Cockayne syndrome. Prenatal diagnosis can be provided only if molecular diagnosis is established in the proband.

The molecular genetics of UV-Sensitive syndrome: A rare dermal anomaly.

Ultraviolet-sensitive syndrome is a rare skin disorder characterised by heterogeneous phenotypic spectrum of skin freckling, telangiectasia and acute sunburn. It usually has an autosomal recessive pattern. So far, only 18 patients from nine different families (Japanese, French, Israeli, Iranian and Pakistani) have been reported in scientific literature. Its precise prevalence is still unknown, but, according to an estimate, its prevalence ratio is 1:100,000 worldwide. Until now, only three genes have been reported to be involved in the syndrome; the Excision Repair Cross-Complementing, Group 6, the Excision Repair Cross-Complementing, Group 8 and the UV-Stimulated Scaffold Protein A (UVSSA). Among these genes, the last one is reported to be more prevalent among different ethnicities, including Pakistani. Physiologically, most of the syndrome genes are involved in the transcription-coupled nucleotide excision pathway. In order to reduce the disease severity, the patients are advised to use medicated skin moisturisers or sun-blocks, sunglasses and gloves, while going out in the sun to avoid sun exposure. The current narrative review was planned to discuss the molecular genetics and the mutational spectrum of the syndrome, and to describe the differential diagnosis of various related disorders in order to facilitate clinical researchers.

Atypical features and de novo heterozygous mutations in two siblings with Cockayne syndrome.

BACKGROUND: Cockayne syndrome (CS) is a rare autosomal recessive disorder which displays multiorgan dysfunction, especially within the nervous system including psychomotor retardation, cerebral atrophy, microcephaly, cognitive dysfunction, mental retardation, and seizures. Many genetic variations reported were related to this syndrome, but splicing mutations with cardiac anomalies have not been found in previous studies. METHODS: Herein, we described a pair of brothers and sisters who present essential manifestations of CS including premature feature, developmental delay, growth failure, microcephaly, and characteristic facial features, such as sunken eyes and a beaked nose. Interestingly, the brother also presented with atypical features which included cardiac anomalies such as left atrioventricular enlargement and cardiac dysfunction such as dilated cardiomyopathy. In addition, whole exome sequencing and RNA sequencing were employed to analyze their genetic landscape. RESULTS: WES analysis showed that these two cases carried double unreported heterozygous spliced mutations in the excision repair cross-complementing group 8 (ERCC8, also known as CSA, NM_000082) gene, which were c.78-2 (IVS1) A>T and c.1042-1 (IVS10) G>A, respectively. Moreover, transcript sequencing analysis validated these mutation sites. In this study, Gene Ontology enrichment and KEGG pathway analyses from RNA sequencing demonstrated similarities but some differences when compared with previous studies. CONCLUSION: For patients with Cockayne syndrome, cardiac changes need to be monitored carefully, especially for cases with splicing mutations of the ERCC8 gene.

Multimodal imaging in a family with Cockayne syndrome with a novel pathogenic mutation in the ERCC8 gene, and significant phenotypic variability.

BACKGROUND: Cockayne syndrome is a rare autosomal recessive neurodegenerative disorder caused by mutations of either the ERCC6/CSB or ERCC8/CSA genes. Here, we describe two sisters with Cockayne syndrome caused by compound heterozygous mutations in the ERCC8 gene using multimodal imaging. Significant ophthalmic and systemic phenotypic variability is discussed. MATERIALS AND METHODS: Multimodal imaging was performed in two affected sisters and included electroretinography, optical coherence tomography, ultra-wide-field confocal scanning laser ophthalmoscopy, fundus autofluorescence and fluorescein angiography, and magnetic resonance imaging. Genetic analyses were performed on the affected sisters, both parents, and three unaffected siblings. RESULTS: The older sister (Patient 1) had mental retardation, bilateral hearing loss, ataxia, and decreased visual acuity with retinal dystrophy. Radiographic studies revealed microcephaly, cerebral and cerebellar atrophy, ventriculomegaly, and a diffusely thickened skull. Full-field electroretinography waveforms were severely diminished with attenuation of cone and rod responses. The younger sister (Patient 2) had similar clinical features, including ataxia, bilateral hearing loss, and decreased visual acuity with retinal dystrophy. She also had paranoid schizophrenia. Wide-field fundus autofluorescence showed scattered areas of retinal pigment epithelium atrophy, which was different from her sister. Genetic analysis revealed two mutations in the ERCC8 gene shared by the sisters. These include an unreported missense point mutation: p.Thr328Ser:c.983C > G, and another previously reported pathogenic missense mutation: p.Ala205Pro:c.613G > C. Familial testing showed in trans segregation of these mutations with unaffected siblings inheriting one or neither mutation, but not both. CONCLUSION: The clinical presentation and genetic studies confirmed a diagnosis of Cockayne syndrome in both sisters caused by compound heterozygous mutations in the ERCC8 gene on chromosome 10. Multimodal ocular imaging and systemic findings revealed wide phenotypic variability between the affected siblings.

Homozygosity mapping and whole exome sequencing reveal a novel ERCC8 mutation in a Chinese consanguineous family with unique cerebellar ataxia.

BACKGROUND: A consanguineous Chinese family was affected by an apparently novel autosomal recessive disorder characterized by cerebellar ataxia, cutaneous photosensitivity, and mild intellectual disability. METHODS: The family was evaluated by homozygosity mapping, haplotype analysis, whole exome sequencing, and candidate gene mutation screening to identify the disease-associated gene and mutation. Bioinformatics methods were used to predict the functional significance of the mutated gene product. ERCC8 mutations and phenotypes were examined. RESULTS: All three patients presented cerebellar ataxia, cutaneous photosensitivity, and mild intellectual disability. Whole genome and candidate region linkage analysis in the consanguineous family revealed a maximum logarithm of the odds score at 5q12.1. This homozygous region was confirmed by homozygosity mapping. The pathogenic missense mutation p.Gly257Arg affecting an evolutionary highly conserved amino acid was identified in ERCC8 at 5q12.1. Integrated application of whole exome sequencing and homozygosity mapping is an efficient approach for gene mapping and mutation identification in consanguineous families. CONCLUSIONS: We identified a novel ERCC8 mutation and new unique disease phenotype. These results also confirmed the genotype-phenotype relationship between mutations in ERCC8 and clinical findings.

First molecular study in Lebanese patients with Cockayne syndrome and report of a novel mutation in ERCC8 gene.

BACKGROUND: Cockayne Syndrome (CS) is a rare autosomal recessive disorder characterized by neurological and sensorial impairment, dwarfism, microcephaly and photosensitivity. CS is caused by mutations in ERCC6 (CSB) or ERCC8 (CSA) genes. METHODS: Three patients with CS were referred to the Medical Genetics Unit of Saint Joseph University. Sanger sequencing of both ERCC8 and ERCC6 genes was performed: ERCC8 was tested in all patients while ERCC6 in one of them. RESULTS: Sequencing led to the identification of three homozygous mutations, two in ERCC8 (p.Y322* and c.843 + 1G > C) and one in ERCC6 (p.R670W). All mutations were previously reported as pathogenic except for the c.843 + 1G > C splice site mutation in ERCC8 which is novel. CONCLUSIONS: Molecular diagnosis was established in all patients included in our study. A genotype-phenotype correlation is discussed and a link, between mutations and some specific religious communities in Lebanon, is suggested.

Exome sequencing revealed a novel deletion in the ERCC8 gene in an Iranian family with Cockayne syndrome.

Cockayne syndrome (CS) is one the rare DNA-repair deficiency disorders with autosomal recessive inheritance. Failure to thrive and microcephaly are the major criteria of diagnosis. Owing to genetic heterogeneity of CS, whole exome sequencing is promising way to determine the genetic basis of the disease. Here, we present c.1053delT in ERCC8 gene in an Iranian family with symptom of CS using whole exome sequencing. The deletion was novel and was not previously reported elsewhere.

Functional and clinical relevance of novel mutations in a large cohort of patients with Cockayne syndrome.

BACKGROUND: Cockayne syndrome (CS) is a rare, autosomal recessive multisystem disorder characterised by prenatal or postnatal growth failure, progressive neurological dysfunction, ocular and skeletal abnormalities and premature ageing. About half of the patients with symptoms diagnostic for CS show cutaneous photosensitivity and an abnormal cellular response to UV light due to mutations in either the ERCC8/CSA or ERCC6/CSB gene. Studies performed thus far have failed to delineate clear genotype-phenotype relationships. We have carried out a four-centre clinical, molecular and cellular analysis of 124 patients with CS. METHODS AND RESULTS: We assigned 39 patients to the ERCC8/CSA and 85 to the ERCC6/CSB genes. Most of the genetic variants were truncations. The missense variants were distributed non-randomly with concentrations in relatively short regions of the respective proteins. Our analyses revealed several hotspots and founder mutations in ERCC6/CSB. Although no unequivocal genotype-phenotype relationships could be made, patients were more likely to have severe clinical features if the mutation was downstream of the PiggyBac insertion in intron 5 of ERCC6/CSB than if it was upstream. Also a higher proportion of severely affected patients was found with mutations in ERCC6/CSB than in ERCC8/CSA. CONCLUSION: By identifying >70 novel homozygous or compound heterozygous genetic variants in 124 patients with CS with different disease severity and ethnic backgrounds, we considerably broaden the CSA and CSB mutation spectrum responsible for CS. Besides providing information relevant for diagnosis of and genetic counselling for this devastating disorder, this study improves the definition of the puzzling genotype-phenotype relationships in patients with CS.

A Novel Mutation in ERCC8 Gene Causing Cockayne Syndrome.

Cockayne syndrome (CS) is a rare autosomal recessive multisystem disorder characterized by impaired neurological and sensory functions, cachectic dwarfism, microcephaly, and photosensitivity. This syndrome shows a variable age of onset and rate of progression, and its phenotypic spectrum include a wide range of severity. Due to the progressive nature of this disorder, diagnosis can be more important when additional signs and symptoms appear gradually and become steadily worse over time. Therefore, mutation analysis of genes involved in CS pathogenesis can be helpful to confirm the suspected clinical diagnosis. Here, we report a novel mutation in ERCC8 gene in a 16-year-old boy who suffers from poor weight gain, short stature, microcephaly, intellectual disability, and photosensitivity. The patient was born to consanguineous family with no previous documented disease in his parents. To identify disease-causing mutation in the patient, whole exome sequencing utilizing next-generation sequencing on an Illumina HiSeq 2000 platform was performed. Results revealed a novel homozygote mutation in ERCC8 gene (NM_000082: exon 11, c.1122G>C) in our patient. Another gene (ERCC6), which is also involved in CS did not have any disease-causing mutations in the proband. The new identified mutation was then confirmed by Sanger sequencing in the proband, his parents, and extended family members, confirming co-segregation with the disease. In addition, different bioinformatics programs which included MutationTaster, I-Mutant v2.0, NNSplice, Combined Annotation Dependent Depletion, The PhastCons, Genomic Evolutationary Rate Profiling conservation score, and T-Coffee Multiple Sequence Alignment predicted the pathogenicity of the mutation. Our study identified a rare novel mutation in ERCC8 gene and help to provide accurate genetic counseling and prenatal diagnosis to minimize new affected individuals in this family.

Epistatic SNP interaction of ERCC6 with ERCC8 and their joint protein expression contribute to gastric cancer/atrophic gastritis risk.

Excision repair cross-complementing group 6 and 8 (ERCC6 and ERCC8) are two indispensable genes for the initiation of transcription-coupled nucleotide excision repair pathway. This study aimed to evaluate the interactions between single nucleotide polymorphisms of ERCC6 (rs1917799) and ERCC8 (rs158572 and rs158916) in gastric cancer and its precancerous diseases. Besides, protein level analysis were performed to compare ERCC6 and ERCC8 expression in different stages of gastric diseases, and to correlate SNPs jointly with gene expression. Sequenom MassARRAY platform method was used to detect polymorphisms of ERCC6 and ERCC8 in 1916 subjects. In situ ERCC6 and ERCC8 protein expression were detected by immunohistochemistry in 109 chronic superficial gastritis, 109 chronic atrophic gastritis and 109 gastric cancer cases. Our results demonstrated pairwise epistatic interactions between ERCC6 and ERCC8 SNPs that ERCC6 rs1917799-ERCC8 rs158572 combination was associated with decreased risk of chronic atrophic gastritis and increased risk of gastric cancer. ERCC6 rs1917799 also showed a significant interaction with ERCC8 rs158916 to reduce gastric cancer risk. The expressions of ERCC6, ERCC8 and ERCC6-ERCC8 combination have similarities that higher positivity was observed in chronic superficial gastritis compared with chronic atrophic gastritis and gastric cancer. As for the effects of ERCC6 and ERCC8 SNPs on the protein expression, single SNP had no correlation with corresponding gene expression, whereas the ERCC6 rs1917799-ERCC8 rs158572 pair had significant influence on ERCC6 and ERCC6-ERCC8 expression. In conclusion, ERCC6 rs1917799, ERCC8 rs158572 and rs158916 demonstrated pairwise epistatic interactions to associate with chronic atrophic gastritis and gastric cancer risk. The ERCC6 rs1917799-ERCC8 rs158572 pair significantly influence ERCC6 and ERCC6-ERCC8 expression.

Uncommon nucleotide excision repair phenotypes revealed by targeted high-throughput sequencing.

BACKGROUND: Deficient nucleotide excision repair (NER) activity causes a variety of autosomal recessive diseases including xeroderma pigmentosum (XP) a disorder which pre-disposes to skin cancer, and the severe multisystem condition known as Cockayne syndrome (CS). In view of the clinical overlap between NER-related disorders, as well as the existence of multiple phenotypes and the numerous genes involved, we developed a new diagnostic approach based on the enrichment of 16 NER-related genes by multiplex amplification coupled with next-generation sequencing (NGS). METHODS: Our test cohort consisted of 11 DNA samples, all with known mutations and/or non pathogenic SNPs in two of the tested genes. We then used the same technique to analyse samples from a prospective cohort of 40 patients. Multiplex amplification and sequencing were performed using AmpliSeq protocol on the Ion Torrent PGM (Life Technologies). RESULTS: We identified causative mutations in 17 out of the 40 patients (43%). Four patients showed biallelic mutations in the ERCC6(CSB) gene, five in the ERCC8(CSA) gene: most of them had classical CS features but some had very mild and incomplete phenotypes. A small cohort of 4 unrelated classic XP patients from the Basque country (Northern Spain) revealed a common splicing mutation in POLH (XP-variant), demonstrating a new founder effect in this population. Interestingly, our results also found ERCC2(XPD), ERCC3(XPB) or ERCC5(XPG) mutations in two cases of UV-sensitive syndrome and in two cases with mixed XP/CS phenotypes. CONCLUSIONS: Our study confirms that NGS is an efficient technique for the analysis of NER-related disorders on a molecular level. It is particularly useful for phenotypes with combined features or unusually mild symptoms. Targeted NGS used in conjunction with DNA repair functional tests and precise clinical evaluation permits rapid and cost-effective diagnosis in patients with NER-defects.