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International Journal of Molecular... May 2024Retinitis pigmentosa (RP) is an inherited retinal dystrophy caused by the loss of photoreceptors and retinal pigment epithelial atrophy, leading to severe visual...
Retinitis pigmentosa (RP) is an inherited retinal dystrophy caused by the loss of photoreceptors and retinal pigment epithelial atrophy, leading to severe visual impairment or blindness. RP can be classified as nonsyndromic or syndromic with complex clinical phenotypes. Three unrelated Polish probands affected with retinitis pigmentosa coexisting with cerebellar ataxia were recruited for this study. Clinical heterogeneity and delayed appearance of typical disease symptoms significantly prolonged the patients' diagnostic process. Therefore, many clinical and genetic tests have been performed in the past. Here, we provide detailed clinical and genetic analysis results of the patients. Whole-exome sequencing (WES) and targeted NGS analysis allow the identification of four novel and two previously reported variants in the following genes: , , and The use of next-generation sequencing (NGS) methods finally allowed for confirmation of the clinical diagnosis. Ultra-rare diseases such as PHARC, PCARP, and Oliver-McFarlane syndromes were diagnosed in patients, respectively. Our findings confirmed the importance of the application of next-generation sequencing methods, especially in ultra-rare genetic disorders with overlapping features.
Topics: Humans; Retinitis Pigmentosa; Male; Female; Exome Sequencing; Pedigree; High-Throughput Nucleotide Sequencing; Adult; Cerebellar Ataxia; Membrane Transport Proteins; Monoacylglycerol Lipases; Mutation; Ataxia; Phenotype; Acyltransferases; Cataract; Phospholipases; Polyneuropathies
PubMed: 38891946
DOI: 10.3390/ijms25115759 -
International Journal of Molecular... May 2024Developmental and epileptic encephalopathy-9 (DEE9) is characterized by seizure onset in infancy, mild to severe intellectual impairment, and psychiatric features and is...
Developmental and epileptic encephalopathy-9 (DEE9) is characterized by seizure onset in infancy, mild to severe intellectual impairment, and psychiatric features and is caused by a mutation in the gene on chromosome Xq22. The rare, unusual X-linked type of disorder affects heterozygous females and mosaic males; transmitting males are unaffected. In our study, 165 patients with epilepsy were tested by Next Generation Sequencing (NGS)-based panel and exome sequencing using Illumina technology. screening identified three point mutations, one indel, and one 29 bp-long deletion in five unrelated female probands. Two novel mutations, c.1152_1180del (p.Gln385Serfs*6) and c.830_831delinsAA (p.Phe277*), were identified and found to be de novo pathogenic. Moreover, among the three inherited mutations, two originated from asymptomatic mothers and one from an affected father. The c.1682C>T and c.1711G>T mutations were present in the DNA samples of asymptomatic mothers. After targeted parental testing, X chromosome inactivation tests and Sanger sequencing were carried out for mosaicism examination on maternal saliva samples in the two asymptomatic mutation carrier subjects. Tissue mosaicism and X-inactivation tests were negative. Our results support the opportunity for reduced penetrance in DEE9 and contribute to expanding the genotype-phenotype spectrum of -related epilepsy.
Topics: Humans; Protocadherins; Female; Cadherins; Epilepsy; High-Throughput Nucleotide Sequencing; Mutation; Pedigree; Male; Child, Preschool; Child; Infant; Age of Onset
PubMed: 38891919
DOI: 10.3390/ijms25115732 -
Cells May 2024The identification of anticancer therapies using next-generation sequencing (NGS) is necessary for the treatment of cholangiocarcinoma. NGS can be easily performed when...
The identification of anticancer therapies using next-generation sequencing (NGS) is necessary for the treatment of cholangiocarcinoma. NGS can be easily performed when cell blocks (CB) are obtained from bile stored overnight. We compared NGS results of paired CB and surgically resected specimens (SRS) from the same cholangiocarcinoma cases. Of the prospectively collected 64 bile CBs from 2018 to 2023, NGS was performed for three cases of cholangiocarcinoma that could be compared with the SRS results. The median numbers of DNA and RNA reads were 95,077,806 [CB] vs. 93,161,788 [SRS] and 22,101,328 [CB] vs. 24,806,180 [SRS], respectively. We evaluated 588 genes and found that almost all genetic alterations were attributed to single-nucleotide variants, insertions/deletions, and multi-nucleotide variants. The coverage rate of variants in SRS by those found in CB was 97.9-99.2%, and the coverage rate of SRS genes by CB genes was 99.6-99.7%. The NGS results of CB fully covered the variants and genetic alterations observed in paired SRS samples. As bile CB is easy to prepare in general hospitals, our results suggest the potential use of bile CB as a novel method for NGS-based evaluation of cholangiocarcinoma.
Topics: Cholangiocarcinoma; Humans; High-Throughput Nucleotide Sequencing; Bile; Male; Middle Aged; Female; Bile Duct Neoplasms; Aged; Mutation
PubMed: 38891057
DOI: 10.3390/cells13110925 -
Scientific Reports Jun 2024The composition of cell-type is a key indicator of health. Advancements in bulk gene expression data curation, single cell RNA-sequencing technologies, and computational...
The composition of cell-type is a key indicator of health. Advancements in bulk gene expression data curation, single cell RNA-sequencing technologies, and computational deconvolution approaches offer a new perspective to learn about the composition of different cell types in a quick and affordable way. In this study, we developed a quantile regression and deep learning-based method called Neural Network Immune Contexture Estimator (NNICE) to estimate the cell type abundance and its uncertainty by automatically deconvolving bulk RNA-seq data. The proposed NNICE model was able to successfully recover ground-truth cell type fraction values given unseen bulk mixture gene expression profiles from the same dataset it was trained on. Compared with baseline methods, NNICE achieved better performance on deconvolve both pseudo-bulk gene expressions (Pearson correlation R = 0.9) and real bulk gene expression data (Pearson correlation R = 0.9) across all cell types. In conclusion, NNICE combines statistic inference with deep learning to provide accurate and interpretable cell type deconvolution from bulk gene expression.
Topics: Neural Networks, Computer; Deep Learning; Humans; Algorithms; Gene Expression Profiling; Single-Cell Analysis; Computational Biology; RNA-Seq; Sequence Analysis, RNA; Transcriptome
PubMed: 38890415
DOI: 10.1038/s41598-024-65053-w -
Scientific Reports Jun 2024Canine liposarcoma is an uncommon tumor that shares morphological similarities with its human counterpart. In dogs, the genetic features of this tumor are unknown and,...
Canine liposarcoma is an uncommon tumor that shares morphological similarities with its human counterpart. In dogs, the genetic features of this tumor are unknown and, based on immunohistochemical studies, amplification of the gene MDM2 and the mutation of TP53 are suspected. In this study 51 cases of primary liposarcomas were immunohistochemically stained for MDM2 and p53 and subjected to fluorescent in situ hybridization and next-generation sequencing to detect MDM2 amplification and TP53 mutations, respectively. MDM2 and p53 were expressed in 21 and 6 cases, respectively. MDM2 amplification and TP53 mutations were identified in 10 and 15 cases, respectively. Statistical analysis revealed an association of the myxoid subtype and the mitotic count with p53 expression and TP53 mutation. No association was found between MDM2 amplification and MDM2 expression or tumor subtype. These results suggest that despite morphological similarities, canine liposarcoma differs from its human counterpart, for which MDM2 amplification is diagnostic for well differentiated and de-differentiated variants, and TP53 mutations are more common in pleomorphic liposarcoma rather than the myxoid one as occur in our cases. Furthermore, canine myxoid liposarcoma likely represents a distinct disease rather than a mere morphological variant.
Topics: Proto-Oncogene Proteins c-mdm2; Dogs; Animals; Liposarcoma; Tumor Suppressor Protein p53; Dog Diseases; Mutation; Female; Male; In Situ Hybridization, Fluorescence; High-Throughput Nucleotide Sequencing; Gene Amplification; Immunohistochemistry
PubMed: 38890407
DOI: 10.1038/s41598-024-64963-z -
Scientific Reports Jun 2024Programmed Death Receptor 1 (PD-1) inhibitors, when combined with chemotherapy, have exhibited notable effectiveness in enhancing the survival outcomes of patients...
Programmed Death Receptor 1 (PD-1) inhibitors, when combined with chemotherapy, have exhibited notable effectiveness in enhancing the survival outcomes of patients afflicted with advanced gastric cancer. However, it is important to acknowledge that not all patients derive substantial benefits from this therapeutic approach, highlighting the crucial necessity of identifying efficacious biomarkers to inform immunotherapy interventions. In this study, we sought to investigate the predictive utility of circulating tumor DNA (ctDNA) as a biomarker in a cohort of 30 patients diagnosed with advanced gastric cancer, all of whom underwent first-line treatment involving PD-1 inhibitor administration alongside chemotherapy. We procured peripheral blood samples both at baseline and following the completion of two treatment cycles. Additionally, baseline tissue specimens were collected for the purpose of genomic alteration assessment, employing both 47-gene and 737-gene next-generation sequencing panels for plasma and tumor tissue, respectively. We delineated a ctDNA response as the eradication of maximum variant allele frequencies relative to baseline levels. Notably, the objective response rate among individuals exhibiting a ctDNA response proved significantly superior in comparison to non-responders (P = 0.0073). Furthermore, patients who manifested a ctDNA response experienced markedly prolonged progression-free survival (PFS) and overall survival (OS) when juxtaposed with those devoid of a ctDNA response (median PFS: 15.6 vs. 6.0 months, P = 0.003; median OS: not reached [NR] vs. 9.0 months, P = 0.011). In summation, patients with advanced gastric cancer receiving first-line treatment with PD-1 inhibitors and chemotherapy, dynamic changes in ctDNA can serve as a potential biomarker for predicting treatment efficacy and long-term outcomes.
Topics: Humans; Stomach Neoplasms; Circulating Tumor DNA; Male; Female; Middle Aged; Aged; Immune Checkpoint Inhibitors; Biomarkers, Tumor; Programmed Cell Death 1 Receptor; Adult; Treatment Outcome; Antineoplastic Combined Chemotherapy Protocols; High-Throughput Nucleotide Sequencing
PubMed: 38890392
DOI: 10.1038/s41598-024-63486-x -
Nature Communications Jun 2024Multi-drug resistant (MDR) E. coli constitute a major public health burden globally, reaching the highest prevalence in the global south yet frequently flowing with...
Multi-drug resistant (MDR) E. coli constitute a major public health burden globally, reaching the highest prevalence in the global south yet frequently flowing with travellers to other regions. However, our comprehension of the entire genetic diversity of E. coli colonising local populations remains limited. We quantified this diversity, its associated antimicrobial resistance (AMR), and assessed the impact of antibiotic use by recruiting 494 outpatients and 423 community dwellers in the Punjab province, Pakistan. Rectal swab and stool samples were cultured on CLED agar and DNA extracted from plate sweeps was sequenced en masse to capture both the genetic and AMR diversity of E. coli. We assembled 5,247 E. coli genomes from 1,411 samples, displaying marked genetic diversity in gut colonisation. Compared with high income countries, the Punjabi population generally showed a markedly different distribution of genetic lineages and AMR determinants, while use of antibiotics elevated the prevalence of well-known globally circulating MDR clinical strains. These findings implicate that longitudinal multi-regional genomics-based surveillance of both colonisation and infections is a prerequisite for developing mechanistic understanding of the interplay between ecology and evolution in the maintenance and dissemination of (MDR) E. coli.
Topics: Pakistan; Humans; Escherichia coli; Anti-Bacterial Agents; Escherichia coli Infections; Drug Resistance, Multiple, Bacterial; High-Throughput Nucleotide Sequencing; Feces; Female; Male; Genome, Bacterial; Adult; Genetic Variation; Middle Aged; Young Adult; Phylogeny; Adolescent; Child
PubMed: 38890378
DOI: 10.1038/s41467-024-49591-5 -
Nature Communications Jun 2024Telomeres are the protective nucleoprotein structures at the end of linear eukaryotic chromosomes. Telomeres' repetitive nature and length have traditionally challenged...
Telomeres are the protective nucleoprotein structures at the end of linear eukaryotic chromosomes. Telomeres' repetitive nature and length have traditionally challenged the precise assessment of the composition and length of individual human telomeres. Here, we present Telo-seq to resolve bulk, chromosome arm-specific and allele-specific human telomere lengths using Oxford Nanopore Technologies' native long-read sequencing. Telo-seq resolves telomere shortening in five population doubling increments and reveals intrasample, chromosome arm-specific, allele-specific telomere length heterogeneity. Telo-seq can reliably discriminate between telomerase- and ALT-positive cancer cell lines. Thus, Telo-seq is a tool to study telomere biology during development, aging, and cancer at unprecedented resolution.
Topics: Humans; Telomere; Neoplasms; Aging; Telomerase; Cell Line, Tumor; Telomere Shortening; Sequence Analysis, DNA; High-Throughput Nucleotide Sequencing; Alleles
PubMed: 38890299
DOI: 10.1038/s41467-024-48917-7 -
Clinical and Experimental Medicine Jun 2024To determine the diagnostic yield of Next-generation sequencing (NGS) in suspect Primary Immunodeficiencies Diseases (PIDs). This systematic review was conducted... (Meta-Analysis)
Meta-Analysis Review
To determine the diagnostic yield of Next-generation sequencing (NGS) in suspect Primary Immunodeficiencies Diseases (PIDs). This systematic review was conducted following PRISMA criteria. Searching Pubmed and Web of Science databases, the following keywords were used in the search: ("Next-generation sequencing") OR "whole exome sequencing" OR "whole genome sequencing") AND ("primary immunodeficiency disease" OR "PIDs"). We used STARD items to assess the risk of bias in the included studies. The meta-analysis included 29 studies with 5847 patients, revealing a pooled positive detection rate of 42% (95% CI 0.29-0.54, P < 0.001) for NGS in suspected PID cases. Subgroup analyses based on family history demonstrated a higher detection rate of 58% (95% CI 0.43-0.71) in patients with a family history compared to 33% (95% CI 0.21-0.46) in those without (P < 0.001). Stratification by disease types showed varied detection rates, with Severe Combined Immunodeficiency leading at 58% (P < 0.001). Among 253 PID-related genes, RAG1, ATM, BTK, and others constituted major contributors, with 34 genes not included in the 2022 IUIS gene list. The application of NGS in suspected PID patients can provide significant diagnostic results, especially in patients with a family history. Meanwhile, NGS performs excellently in accurately diagnosing disease types, and early identification of disease types can benefit patients in treatment.
Topics: Humans; High-Throughput Nucleotide Sequencing; Primary Immunodeficiency Diseases
PubMed: 38890201
DOI: 10.1007/s10238-024-01392-2 -
PloS One 2024Single-cell transcriptomics techniques, such as scRNA-seq, attempt to characterize gene expression profiles in each cell of a heterogeneous sample individually. Due to...
Single-cell transcriptomics techniques, such as scRNA-seq, attempt to characterize gene expression profiles in each cell of a heterogeneous sample individually. Due to growing amounts of data generated and the increasing complexity of the computational protocols needed to process the resulting datasets, the demand for dedicated training in mathematical and programming skills may preclude the use of these powerful techniques by many teams. In order to help close that gap between wet-lab and dry-lab capabilities we have developed SinglePointRNA, a shiny-based R application that provides a graphic interface for different publicly available tools to analyze single cell RNA-seq data. The aim of SinglePointRNA is to provide an accessible and transparent tool set to researchers that allows them to perform detailed and custom analysis of their data autonomously. SinglePointRNA is structured in a context-driven framework that prioritizes providing the user with solid qualitative guidance at each step of the analysis process and interpretation of the results. Additionally, the rich user guides accompanying the software are intended to serve as a point of entry for users to learn more about computational techniques applied to single cell data analysis. The SinglePointRNA app, as well as case datasets for the different tutorials are available at www.github.com/ScienceParkMadrid/SinglePointRNA.
Topics: Software; Single-Cell Analysis; RNA-Seq; Humans; Computational Biology; Sequence Analysis, RNA; User-Computer Interface; Gene Expression Profiling
PubMed: 38889133
DOI: 10.1371/journal.pone.0300567