Frequency Of Copy Number Variations Research Articles

The yield of SNP microarray analysis for fetal ultrasound cardiac abnormalities.

Chromosomal microarray analysis (CMA) has emerged as a critical instrument in prenatal diagnostic procedures, notably in assessing congenital heart diseases (CHD). Nonetheless, current research focuses solely on CHD, overlooking the necessity for thorough comparative investigations encompassing fetuses with varied structural abnormalities or those without apparent structural anomalies. This study sought to assess the relation of single nucleotide polymorphism-based chromosomal microarray analysis (SNP-based CMA) in identifying the underlying causes of fetal cardiac ultrasound abnormalities. A total of 2092 pregnant women who underwent prenatal diagnosis from 2017 to 2022 were included in the study and divided into four groups based on the presence of ultrasound structural abnormalities and the specific type of abnormality. The results of the SNP-Array test conducted on amniotic fluid samples from these groups were analyzed. Findings from the study revealed that the non-isolated CHD group exhibited the highest incidence of aneuploidy, overall chromosomal abnormalities, and trisomy 18, demonstrating statistically significant differences from the other groups (p < 0.001). Regarding the distribution frequency of copy number variation (CNV) segment size, no statistically significant distinctions were observed between the isolated CHD group and the non-isolated CHD group (p > 0.05). The occurrence rates of 22q11.2 and 15q11.2 were also not statistically different between the isolated CHD group and the non-isolated congenital heart defect group (p > 0.05). SNP-based CMA enhances the capacity to detect abnormal CNVs in CHD fetuses, offering valuable insights for diagnosing chromosomal etiology and facilitating genetic counseling. This research contributes to the broader understanding of the utility of SNP-based CMA in the context of fetal cardiac ultrasound abnormalities.

Systematic molecular analyses for 115 karyotypically normal men with isolated non-obstructive azoospermia.

Do copy-number variations (CNVs) in the azoospermia factor (AZF) regions and monogenic mutations play a major role in the development of isolated (non-syndromic) non-obstructive azoospermia (NOA) in Japanese men with a normal 46, XY karyotype? Deleterious CNVs in the AZF regions and damaging sequence variants in eight genes likely constitute at least 8% and approximately 8% of the genetic causes, respectively, while variants in other genes play only a minor role. Sex chromosomal abnormalities, AZF-linked microdeletions, and monogenic mutations have been implicated in isolated NOA. More than 160 genes have been reported as causative/susceptibility/candidate genes for NOA. Systematic molecular analyses were conducted for 115 patients with isolated NOA and a normal 46, XY karyotype, who visited our hospital between 2017 and 2021. We studied 115 unrelated Japanese patients. AZF-linked CNVs were examined using sequence-tagged PCR and multiplex ligation-dependent probe amplification, and nucleotide variants were screened using whole exome sequencing (WES). An optimized sequence kernel association test (SKAT-O), a gene-based association study using WES data, was performed to identify novel disease-associated genes in the genome. The results were compared to those of previous studies and our in-house control data. Thirteen types of AZF-linked CNVs, including the hitherto unreported gr/gr triplication and partial AZFb deletion, were identified in 63 (54.8%) cases. When the gr/gr deletion, a common polymorphism in Japan, was excluded from data analyses, the total frequency of CNVs was 23/75 (30.7%). This frequency is higher than that of the reference data in Japan and China (11.1% and 14.7%, respectively). Known NOA-causative AZF-linked CNVs were found in nine (7.8%) cases. Rare damaging variants in known causative genes (DMRT1, PLK4, SYCP2, TEX11, and USP26) and hemizygous/multiple-heterozygous damaging variants in known spermatogenesis-associated genes (TAF7L, DNAH2, and DNAH17) were identified in nine cases (7.8% in total). Some patients carried rare damaging variants in multiple genes. SKAT-O detected no genes whose rare damaging variants were significantly accumulated in the patient group. The number of participants was relatively small, and the clinical information of each patient was fragmentary. Moreover, the pathogenicity of identified variants was assessed only by in silico analyses. This study showed that various AZF-linked CNVs are present in more than half of Japanese NOA patients. These results broadened the structural variations of AZF-linked CNVs, which should be considered for the molecular diagnosis of spermatogenic failure. Furthermore, the results of this study highlight the etiological heterogeneity and possible oligogenicity of isolated NOA. This study was supported by Grants from the Japan Society for the Promotion of Science (21K19283 and 21H0246), the Japan Agency for Medical Research and Development (22ek0109464h0003), the National Center for Child Health and Development, the Canon Foundation, the Japan Endocrine Society, and the Takeda Science Foundation. The results of this study were based on samples and patient data obtained from the International Center for Reproductive Medicine, Dokkyo Medical University Saitama Medical Center, Koshigaya, Japan. The authors have no conflicts of interest to disclose. N/A.

Rare copy number variation in autoimmune Addison's disease.

Autoimmune Addison's disease (AAD) is a rare but life-threatening endocrine disorder caused by an autoimmune destruction of the adrenal cortex. A previous genome-wide association study (GWAS) has shown that common variants near immune-related genes, which mostly encode proteins participating in the immune response, affect the risk of developing this condition. However, little is known about the contribution of copy number variations (CNVs) to AAD susceptibility. We used the genome-wide genotyping data from Norwegian and Swedish individuals (1,182 cases and 3,810 controls) to investigate the putative role of CNVs in the AAD aetiology. Although the frequency of rare CNVs was similar between cases and controls, we observed that larger deletions (>1,000 kb) were more common among patients (OR = 4.23, 95% CI 1.85-9.66, p = 0.0002). Despite this, none of the large case-deletions were conclusively pathogenic, and the clinical presentation and an AAD-polygenic risk score were similar between cases with and without the large CNVs. Among deletions exclusive to individuals with AAD, we highlight two ultra-rare deletions in the genes LRBA and BCL2L11, which we speculate might have contributed to the polygenic risk in these carriers. In conclusion, rare CNVs do not appear to be a major cause of AAD but further studies are needed to ascertain the potential contribution of rare deletions to the polygenic load of AAD susceptibility.

Genetics of inborn errors of immunity: Diagnostic strategies and new approaches to CNV detection.

Genetic diagnosis of inborn errors of immunity (IEI) is complex due to the large number of genes involved and their molecular features. Missense variants have been reported as the most common cause of IEI. However, the frequency of copy number variants (CNVs) may be underestimated since their detection requires specific quantitative techniques. At this point, the use of Next Generation Sequencing (NGS) is acquiring relevance. In this article, we present our experience in the genetic diagnosis of IEI based on three diagnostic algorithms that allowed the detection of single nucleotide variants (SNVs) and CNVs. Following this approximation, 703 index cases were evaluated between 2014 and 2021. Sanger sequencing, MLPA, CGH array, breakpoint spanning PCR or a customized NGS-based multigene-targeted panel were performed. A genetic diagnosis was reached in 142 of the 703 index cases (20%), 19 of them presented deletions as causal variants. Deletions were also detected in 5 affected relatives and 16 healthy carriers during the family studies. Additionally, we compile, characterize and present all the CNVs detected by our diagnostic algorithms, representing the largest cohort of deletions related to IEI to date. Furthermore, three bioinformatic tools (LACONv, XHMM, VarSeq™) based on NGS data were evaluated. VarSeq™ was the most sensitive and specific bioinformatic tool; detecting 21/23 (91%) deletions located in captured regions. Based on our results, we propose a strategy to guide the molecular diagnosis that can be followed by expert and non-expert centres in the field of IEI.

Experience of copy number variation sequencing applied in spontaneous abortion

PurposeWe evaluated the value of copy number variation sequencing (CNV-seq) and quantitative fluorescence (QF)-PCR for analyzing chromosomal abnormalities (CA) in spontaneous abortion specimens.MethodsA total of 650 products of conception (POCs) were collected from spontaneous abortion between April 2018 and May 2020. CNV-seq and QF-PCR were performed to determine the characteristics and frequencies of copy number variants (CNVs) with clinical significance. The clinical features of the patients were recorded.ResultsClinically significant chromosomal abnormalities were identified in 355 (54.6%) POCs, of which 217 (33.4%) were autosomal trisomies, 42(6.5%) were chromosomal monosomies and 40 (6.2%) were pathogenic CNVs (pCNVs). Chromosomal trisomy occurs mainly on chromosomes 15, 16, 18, 21and 22. Monosomy X was not associated with the maternal or gestational age. The frequency of chromosomal abnormalities in miscarriages from women with a normal live birth history was 55.3%; it was 54.4% from women without a normal live birth history (P > 0.05). There were no significant differences among women without, with 1, and with ≥ 2 previous miscarriages regarding the rate of chromosomal abnormalities (P > 0.05); CNVs were less frequently detected in women with advanced maternal age than in women aged ≤ 29 and 30–34 years (P < 0.05).ConclusionChromosomal abnormalities are the most common cause of pregnancy loss, and maternal and gestational ages are strongly associated with fetal autosomal trisomy aberrations. Embryo chromosomal examination is recommended regardless of the gestational age, modes of conception or previous abortion status.

Copy-number variants in the ACMG secondary finding genes: A reporting framework for clinical cytogeneticists

PurposeTo determine the pathogenicity and frequency of copy-number variants (CNV) in the 81 secondary finding (SFv3.2) genes recommended by the American College of Medical Genetics and Genomics (ACMG). MethodsReview of published evidence on pathogenicity of partial or complete copy-number losses or gains in ACMG SFv3.2 was performed. Frequency of reportable CNVs in the ACMG SFv3.2 genes was investigated among 10,959 patients tested by chromosomal microarray analysis in a single academic testing laboratory at the University of Pittsburgh Medical Center during 2011 to 2023. ResultsWe identified 58 ACMG SFv3.2 genes for which sufficient evidence supports reporting of partial or complete copy-number losses as secondary findings. On the contrary, reporting of copy-number gains was not supported by evidence in any of the ACMG SFv3.2 genes. Overall, CNVs in SFv3.2 genes were detected in 32 of 10,959 (0.29% or 1 in 343) patients in our cohort. ConclusionThis study provides a framework for consistent reporting of CNVs, detected by chromosomal microarray analysis, exome, or genome sequencing, in any of the ACMG SFv3.2 genes. To our knowledge, this is the largest cohort of patients studied for estimation of frequency of reportable CNVs in the ACMG SFv3.2 genes.

Pan-cancer analyses of senescence-related genes in extracellular matrix characterization in cancer

PurposeThe aged microenvironment plays a crucial role in tumor onset and progression. However, it remains unclear whether and how the aging of the extracellular matrix (ECM) influences cancer onset and progression. Furthermore, the mechanisms and implications of extracellular matrix senescence-related genes (ECM-SRGs) in pan-cancer have not been investigated.MethodsWe collected profiling data from over 10,000 individuals, covering 33 cancer types, 750 small molecule drugs, and 24 immune cell types, for a thorough and systematic analysis of ECM-SRGs in cancer.ResultsWe observed a significant correlation between immune cell infiltrates and Gene Set Variation Analysis enrichment scores of ECM-SRGs in 33 cancer types. Moreover, our results revealed significant differences in immune cell infiltration among patients with copy number variations (CNV) and single nucleotide variations (SNV) in ECM-SRGs across various malignancies. Aberrant hypomethylation led to increased ECM-SRGs expression, and in specific malignancies, a connection between ECM-SRGs hypomethylation and adverse patient survival was established. The frequency of CNV and SNV in ECM-SRGs was elevated. We observed a positive correlation between CNV, SNV, and ECM-SRGs expression. Furthermore, a correlation was found between the high frequency of CNV and SNV in ECM-SRGs and poor patient survival in several cancer types. Additionally, the results demonstrated that ECM-SRGs expression could serve as a predictor of patient survival in diverse cancers. Pathway analysis unveiled the role of ECM-SRGs in activating EMT, apoptosis, and the RAS/MAPK signaling pathway while suppressing the cell cycle, hormone AR, and the response to DNA damage signaling pathway. Finally, we conducted searches in the “Genomics of Drug Sensitivity in Cancer” and “Genomics of Therapeutics Response Portal” databases, identifying several drugs that target ECM-SRGs.ConclusionsWe conducted a comprehensive evaluation of the genomes and immunogenomics of ECM-SRGs, along with their clinical features in 33 solid tumors. This may provide insights into the relationship between ECM-SRGs and tumorigenesis. Consequently, targeting these ECM-SRGs holds promise as a clinical approach for cancer treatment.

Genetic and Epigenetic Characteristics in Isolated Pancreatic Metastases of Clear-Cell Renal Cell Carcinoma.

Isolated pancreatic metastases of renal cell carcinoma (IsPMRCC) are a rare manifestation of metastatic, clear-cell renal cell carcinoma (RCC) in which distant metastases occur exclusively in the pancreas. In addition to the main symptom of the isolated occurrence of pancreatic metastases, the entity surprises with additional clinical peculiarities: (a) the unusually long interval of about 9 years between the primary RCC and the onset of pancreatic metastases; (b) multiple pancreatic metastases occurring in 36% of cases; (c) favourable treatment outcomes with a 75% 5-year survival rate; and (d) volume and growth-rate dependent risk factors generally accepted to be relevant for overall survival in metastatic surgery are insignificant in isPMRCC. The genetic and epigenetic causes of exclusive pancreatic involvement have not yet been investigated and are currently unknown. Conversely, according to the few available data in the literature, the following genetic and epigenetic peculiarities can already be identified as the cause of the protracted course: 1. high genetic stability of the tumour cell clones in both the primary tumour and the pancreatic metastases; 2. a low frequency of copy number variants associated with aggressiveness, such as 9p, 14q and 4q loss; 3. in the chromatin-modifying genes, a decreased rate of PAB1 (3%) and an increased rate of PBRM1 (77%) defects are seen, a profile associated with a favourable course; 4. an increased incidence of KDM5C mutations, which, in common with increased PBRM1 alterations, is also associated with a favourable outcome; and 5. angiogenetic biomarkers are increased in tumour tissue, while inflammatory biomarkers are decreased, which explains the good response to TKI therapy and lack of sensitivity to IT.

Genome-wide copy number variations in Bhutia equine breed using SNP genotyping data

Copy number variants (CNVs) have dynamic potential and evolutionary significance like other genetic variants, namely, single nucleotide polymorphisms, InDels, short tandem repeat polymorphisms, inversion variants, etc. Discovering CNVs leads to further speculation that the genomic DNA contains more changes than previously thought and contributes to the phenotypic variation. CNVs are big DNA fragments (&gt; 1 kb) being duplicated or deleted. A bridge between CNVs and phenotypic variations supports CNVs to be utilized in GWAS, which are currently mostly based on SNPs. CNV, which refers to the structural differences, influence gene expression and can be an indicator of numerous traits for improvement. There is a severe dearth of research on CNVs in animals, especially equine. The present study investigates the genomes of the Bhutia Equine breed for genome-wide discovery of CNVs using theAxiom™ Equine Genotyping Array chip for a better understanding of its traits which had been unexplored till date. A total of 619 CNVs from 20 Bhutia equines were identified with the median and average size as 49.394 kb and 114.955 kb, respectively. Total 225 frequent CNVRs with &gt; 1% CNV frequency were identified among them along with singleton type. These CNVRs contained 361 genes in all. The information obtained on genomic variation could be utilized to identify economically advantageous genetic features in Bhutia equine breed.

Abnormal genetic and epigenetic patterns of m6A regulators associated with tumor microenvironment in colorectal cancer.

N6-methyladenosine (m6A) has a critical role in the development and progression of cancer. However, the genetic and epigenetic patterns, as well as tumor microenvironment (TME) infiltration characteristics of m6A regulators in colorectal cancer (CRC) remain largely unknown. Molecular patterns of m6A modifications of 24 m6A regulators in CRC samples were evaluated using data from The Cancer Genome Atlas (TCGA). Mutations, copy number variations (CNVs), DNA methylation, and chromatin accessibility were examined to investigate the underlying mechanisms of the aberrant expression of m6A regulators. Correlations between m6A-related genes and TME cell-infiltrating characteristics were evaluated using Tumor Immune Estimation Resource (TIMER). The m6A regulators were frequently dysregulated in CRC, with two downregulated and 16 upregulated. All the m6A regulators had mutations (frequency ranging from 0.9% to 7%), with active mutations tending to occur in RBM15 and inactive mutations in ZC3H13. Only five m6A regulators had CNV frequency greater than 1%: YTHDC2 (2.4%), YTHDF1 (7.0%), YTHDF3 (1.9%), VIRMA (1.7%), and ZC3H13 (3.0%). The copy numbers of these five genes were positively correlated with their expression levels. The m6A regulators frequently showed imbalanced methylation in CRC, with hypomethylation of YTHDF2, IGF2BP3, FTO, and hypermethylation of HNRNPC, METTL3, and WTAP. Most m6A regulators had high chromatin accessibility, which was positively correlated with their gene expression. IGF2BP1 was identified as an independent prognostic factor for overall survival. Moreover, the expression of most m6A regulators was positively correlated with the infiltration of B cells, CD8+ T cells, CD4+ T cells, macrophages, neutrophils, and dendritic cells. Aberrant expression of m6A regulators is associated with mutation, CNV, and chromatin accessibility, owing to both genetic and epigenetic modifications. The TME infiltration characterization of m6A regulators could guide the development of more effective immunotherapy strategies in CRC.

Mapping the tumor microenvironment in clear cell renal carcinoma by single-cell transcriptome analysis.

Introduction: Clear cell renal cell carcinoma (ccRCC) is associated with unfavorable clinical outcomes. To identify viable therapeutic targets, a comprehensive understanding of intratumoral heterogeneity is crucial. In this study, we conducted bioinformatic analysis to scrutinize single-cell RNA sequencing data of ccRCC tumor and para-tumor samples, aiming to elucidate the intratumoral heterogeneity in the ccRCC tumor microenvironment (TME). Methods: A total of 51,780 single cells from seven ccRCC tumors and five para-tumor samples were identified and grouped into 11 cell lineages using bioinformatic analysis. These lineages included tumor cells, myeloid cells, T-cells, fibroblasts, and endothelial cells, indicating a high degree of heterogeneity in the TME. Copy number variation (CNV) analysis was performed to compare CNV frequencies between tumor and normal cells. The myeloid cell population was further re-clustered into three major subgroups: monocytes, macrophages, and dendritic cells. Differential expression analysis, gene ontology, and gene set enrichment analysis were employed to assess inter-cluster and intra-cluster functional heterogeneity within the ccRCC TME. Results: Our findings revealed that immune cells in the TME predominantly adopted an inflammatory suppression state, promoting tumor cell growth and immune evasion. Additionally, tumor cells exhibited higher CNV frequencies compared to normal cells. The myeloid cell subgroups demonstrated distinct functional properties, with monocytes, macrophages, and dendritic cells displaying diverse roles in the TME. Certain immune cells exhibited pro-tumor and immunosuppressive effects, while others demonstrated antitumor and immunostimulatory properties. Conclusion: This study contributes to the understanding of intratumoral heterogeneity in the ccRCC TME and provides potential therapeutic targets for ccRCC treatment. The findings emphasize the importance of considering the diverse functional roles of immune cells in the TME for effective therapeutic interventions.

Identification of copy number variants with genome sequencing: Clinical experiences from the NYCKidSeq program.

Copy number variations (CNVs) play a significant role in human disease. While chromosomal microarray has traditionally been the first-tier test for CNV detection, use of genome sequencing (GS) is increasing. We report the frequency of CNVs detected with GS in a diverse pediatric cohort from the NYCKidSeq program and highlight specific examples of its clinical impact. A total of 1052 children (0-21 years) with neurodevelopmental, cardiac, and/or immunodeficiency phenotypes received GS. Phenotype-driven analysis was used, resulting in 183 (17.4%) participants with a diagnostic result. CNVs accounted for 20.2% of participants with a diagnostic result (37/183) and ranged from 0.5 kb to 16 Mb. Of participants with a diagnostic result (n = 183) and phenotypes in more than one category, 5/17 (29.4%) were solved by a CNV finding, suggesting a high prevalence of diagnostic CNVs in participants with complex phenotypes. Thirteen participants with a diagnostic CNV (35.1%) had previously uninformative genetic testing, of which nine included a chromosomal microarray. This study demonstrates the benefits of GS for reliable detection of CNVs in a pediatric cohort with variable phenotypes.

Genetic factors inherited from both diploid parents interact to affect genome stability and fertility in resynthesized allotetraploid Brassica napus.

Established allopolyploids are known to be genomically stable and fertile. However, in contrast, most newly resynthesized allopolyploids are infertile and meiotically unstable. Identifying the genetic factors responsible for genome stability in newly formed allopolyploid is key to understanding how 2 genomes come together to form a species. One hypothesis is that established allopolyploids may have inherited specific alleles from their diploid progenitors which conferred meiotic stability. Resynthesized Brassica napus lines are often unstable and infertile, unlike B. napus cultivars. We tested this hypothesis by characterizing 41 resynthesized B. napus lines produced by crosses between 8 Brassica rapa and 8 Brassica oleracea lines for copy number variation resulting from nonhomologous recombination events and fertility. We resequenced 8 B. rapa and 5 B. oleracea parent accessions and analyzed 19 resynthesized lines for allelic variation in a list of meiosis gene homologs. SNP genotyping was performed using the Illumina Infinium Brassica 60K array for 3 individuals per line. Self-pollinated seed set and genome stability (number of copy number variants) were significantly affected by the interaction between both B. rapa and B. oleracea parental genotypes. We identified 13 putative meiosis gene candidates which were significantly associated with frequency of copy number variants and which contained putatively harmful mutations in meiosis gene haplotypes for further investigation. Our results support the hypothesis that allelic variants inherited from parental genotypes affect genome stability and fertility in resynthesized rapeseed.

Comprehensive genomic and transcriptomic profiling analysis of the recurrent gastrointestinal stromal tumor (GIST).

e23506 Background: The gastrointestinal stromal tumor (GIST) is a potentially malignant mesenchymal tumor (sarcoma), whose management has been transformed by approvals of TKIs. However, recurrence or metastasis is a formidable challenge in managing high-risk GIST. Comprehensive investigation of the transcriptome and genome in GIST is helpful to determine the therapy strategy for patients. This study aims to explore the molecular features of high-risk recurrence GIST. Methods: The tumor tissue samples of 42 patients with primary GIST were collected, in which all patients were divided into low-risk (n = 12), intermediate-risk (n = 11), and high-risk (n = 19) groups according to the NIH 2017th, in which 8 developed recurrence within three years and 11 without recurrence over three years in the high-risk group. Tumor-normal whole-exome and tumor mRNA sequencing was performed on baseline tissue DNA isolated from all patients. Results: Consistent with prior knowledge, KIT was the predominant mutation. The top mutated genes in GISTs were KIT (82%), PHLDA1 (39%), MUC6 (22%), ARID1B(20%), SURF2(20%), MGA(16%), MUC17(16%), MPRIP(16%) and PDGFRA(16%), which MUC6 was a significantly different mutation gene (DMG) associated with disease progression in terms of either high-risk(p = 0.04), recurrence(p = 0.02) or progression(p = 0.05). Interestingly, PDGFRA mutation co-occurs with CTNND2(p &lt; 0.01) and its correlated genes. Additionally, we further explored the somatic copy number variations (CNVs). amp_RAD51B(49%), amp_MLH3(41%), amp_CHEK2(37%), and amp_RB1(12%) seemed to be more frequent in the recurrence group, while these observations were not significant (p &gt; 0.05). Notably, del_NCOR1 or amp_NCOR1 was uniquely observed in the recurrence group. Compared with non-recurrence GIST, gene expression profiles revealed that mast cell activate (MCA)，M1-macrophages infiltration were higher in recurrence group, and mRNA was significantly upregulated in Systemic lupus erythematosus、Neutrophil extracellular trap formation、Viral carcinogenesis、Endocytosis. These pathways were indentified as being associated with recurrence and progression in other cancer studies. For example, the endocytosis pathway expression is positively associated with oncogenic pathway signaling in CRC, and neutrophil extracellular trap on cancer cells promotes liver metastasis in the tumor microenvironment. In the other, NCOR1 is a crucial regulator of T cell and B cell development. Conclusions: Here, we demonstrated the DNA/RNA profiles of GISTs. We observed higher CNVs in high-risk GIST. Especially, the high-risk recurred tumor presented different CNVs frequency and microenvironment. These results may contribute to understanding the mechanism of recurrence and improving recurrence detection.

Rare recurrent copy number variations in metabotropic glutamate receptor interacting genes in children with neurodevelopmental disorders

BackgroundNeurodevelopmental disorders (NDDs), such as attention deficit hyperactivity disorder (ADHD) and autism spectrum disorder (ASD), are examples of complex and partially overlapping phenotypes that often lack definitive corroborating genetic information. ADHD and ASD have complex genetic associations implicated by rare recurrent copy number variations (CNVs). Both of these NDDs have been shown to share similar biological etiologies as well as genetic pleiotropy.MethodsPlatforms aimed at investigating genetic-based associations, such as high-density microarray technologies, have been groundbreaking techniques in the field of complex diseases, aimed at elucidating the underlying disease biology. Previous studies have uncovered CNVs associated with genes within shared candidate genomic networks, including glutamate receptor genes, across multiple different NDDs. To examine shared biological pathways across two of the most common NDDs, we investigated CNVs across 15,689 individuals with ADHD (n = 7920), ASD (n = 4318), or both (n = 3,416), as well as 19,993 controls. Cases and controls were matched by genotype array (i.e., Illumina array versions). Three case–control association studies each calculated and compared the observed vs. expected frequency of CNVs across individual genes, loci, pathways, and gene networks. Quality control measures of confidence in CNV-calling, prior to association analyses, included visual inspection of genotype and hybridization intensity.ResultsHere, we report results from CNV analysis in search for individual genes, loci, pathways, and gene networks. To extend our previous observations implicating a key role of the metabotropic glutamate receptor (mGluR) network in both ADHD and autism, we exhaustively queried patients with ASD and/or ADHD for CNVs associated with the 273 genomic regions of interest within the mGluR gene network (genes with one or two degrees protein–protein interaction with mGluR 1–8 genes). Among CNVs in mGluR network genes, we uncovered CNTN4 deletions enriched in NDD cases (P = 3.22E − 26, OR = 2.49). Additionally, we uncovered PRLHR deletions in 40 ADHD cases and 12 controls (P = 5.26E − 13, OR = 8.45) as well as clinically diagnostic relevant 22q11.2 duplications and 16p11.2 duplications in 23 ADHD + ASD cases and 9 controls (P = 4.08E − 13, OR = 15.05) and 22q11.2 duplications in 34 ADHD + ASD cases and 51 controls (P = 9.21E − 9, OR = 3.93); those control samples were not with previous 22qDS diagnosis in their EHR records.ConclusionTogether, these results suggest that disruption in neuronal cell-adhesion pathways confers significant risk to NDDs and showcase that rare recurrent CNVs in CNTN4, 22q11.2, and 16p11.2 are overrepresented in NDDs that constitute patients predominantly suffering from ADHD and ASD.Trial registrationClinicalTrials.gov Identifier: NCT02286817 First Posted: 10 November 14, ClinicalTrials.gov Identifier: NCT02777931 first posted: 19 May 2016, ClinicalTrials.gov Identifier: NCT03006367 first posted: 30 December 2016, ClinicalTrials.gov Identifier: NCT02895906 first posted: 12 September 2016.

Copy number variations of cytochrome P450 genes in Kinh Vietnamese.

The cytochrome P450 (CYP450) family is well known as a major group of drug metabolizing enzymes. The polymorphism of CYP450 genes is the main factor having an impact on the interindividual difference in drug response, including drug efficacy and drug safety. The single nucleotide polymorphism (SNPs) of Vietnamese Kinh has been widely studied, but information about the copy number variations (CNVs) of other CYP450 genes is still unknown. To identify the CNV variability of CYP450 in 154 healthy unrelated Kinh Vietnamese, except eCYP2D6, which was previously reported. Multiplex Ligation-Dependent Probe Amplification (MLPA) was applied for determination of copy number of 10 CYP450 genes. Later, PCR or quantitative PCR (qPCR) was used to confirm the detected CNVs in randomly chosen subjects. Of the 154 subjects, along with CYP2D6, 4 other CYP450 genes showed CNVs including duplications (CYP1B1), deletions (CYP2A6 and CYP2C9), and both duplications and deletions (CYP2E1). Among these, CYP2A6 exhibited the greatest frequency of CNVs compared with other CYP450, in which CYP2A6Del accounted for 11%. Meanwhile, allele CYP2E1Del showed the lowest frequency with only 0.3%. The present study provides new insight into CYP450 CNVs in the Kinh Vietnamese cohort. Our data have contributed to genetic profiling of CYP450 CNVs in Vietnam, which would be helpful for facilitating implementation of pharmacogenetics in drug dosing adjustment in Vietnam.

The Glioma-IRE project − Molecular profiling in patients with glioma: steps toward an individualized diagnostic and therapeutic approach

BackgroundThis study aimed to characterize the genetic profile of patients with glioma and discuss the impact of next-generation sequencing in glioma diagnosis and treatment.MethodsBetween 2019 and 2022, we analyzed the genetic profile of 99 patients with glioma through the Oncomine Focus Assay. The assay enables the detection of mutations in 52 driver genes, including single nucleotide variants (SNVs), copy number variants (CNVs), and gene fusions. We also collected and analyzed patients’ clinic characteristics and treatment outcomes.ResultsOver a period of 35 months, 700 patients with glioma followed by our neuro-oncology unit were screened, and 99 were enrolled in the study; most of the patients were excluded for inadequate non-morphological MRI or lack/inadequacy of the tissue samples. Based on our findings, most patients with glioma present mutations, such as SNVs, CNVs or gene fusions. Our data were similar to those reported by The Cancer Genome Atlas Program in terms of frequency of SNVs and CNVs, while we observed more cases of gene fusions. Median overall survival, progression-free survival, and time to progression were significantly lower for patients with grade VI glioblastoma than those with other gliomas. Only four patients were offered a targeted treatment based on the mutation detected; however, only one received treatment, the others could not receive the selected treatment because of worsening clinical status.ConclusionRoutine timely molecular profiling in patients with glioma should be implemented to offer patients an individualized diagnostic approach and provide them with advanced targeted therapy options if available.

Mutation spectrum and frequency of copy number variations of the ANOS1 gene in patients with Kallmann syndrome or normosmic isolated hypogonadotropic hypogonadism.

This study was performed to investigate the molecular characteristics and frequency of copy number variations (CNVs) of ANOS1 in patients with Kallmann syndrome (KS) or normosmic isolated hypogonadotropic hypogonadism (nIHH) using multiplex ligation-dependent probe amplification (MLPA) analysis and sequencing. Among 45 patients from 43 independent families, Sanger sequencing, next-generation sequencing (NGS), or microarray was performed in 24 patients from 23 families, and MLPA was performed in 19 patients who did not show rare sequence variants (n = 18) or ANOS1 amplification by PCR (n = 1). Seven patients (4 patients with KS, 1 patient with nIHH, one prepubertal boy with anosmia, and one newborn patient) from 6 families (6/43, 14%) harbored molecular defects in ANOS1 including a nonsense mutation [c.1140G>A (p.W380*)], a frameshift mutation [c.1260del (p.Q421Kfs*61)], a splice site mutation (c.1449+1G>A), an exon 7 deletion, a complete deletion, and 7.9 Mb-sized inversion encompassing ANOS1. The complete deletion of ANOS1 was identified in a neonate with a micropenis and cryptorchidism. Unilateral renal agenesis was found in three patients, whereas only one patient displayed both synkinesia and sensorineural hearing loss. There was no reversal of hypogonadotropic hypogonadism in any patient during 9.1 ± 2.9 years of treatment with testosterone enanthate. Molecular defects in the ANOS1 gene could be identified in 14% of probands including various types of CNVs (3/43, 7.0%). Comprehensive analysis using sequencing and analysis for CNVs is required to detect molecular defects in ANOS1.

Comprehensive pan-cancer analysis of role of GPRASP1, associated with clinical outcomes, immune microenvironment, and immunotherapeutic efficiency in pancreatic cancer.

GPRASP1 (G-protein-coupled receptor-associated sorting protein 1) plays an important role in tumorigenesis. However, GPRASP1 specific role has not been clearly clarified in cancer, particularly in pancreatic cancer(PC). Firstly, we utilized pan-cancer analysis based on RNA sequencing data from TCGA (The Cancer Genome Atlas) to evaluate the expression pattern and immunological role of GPRASP1. Then, through multiple transcriptome datasets (TCGA and Gene Expression Omnibus (GEO)) and multi-omics (RNA-seq, DNA methylation, copy number variations (CNV), somatic mutation data) in-depth analysis, we comprehensively explore the relationship of GPRASP1 expression with clinicopathologic characteristics, clinical outcomes, CNV, and DNA methylation in pancreatic cancer. Additionally, we employed immunohistochemistry (IHC) to further confirm GPRASP1 expression pattern between PC tissues and paracancerous tissues. Lastly, we systematically associated the GPRASP1 with immunological properties from numerous perspectives, such as immune cell infiltration, immune-related pathways, immune checkpoint inhibitors, immunomodulators, immunogenicity, and immunotherapy. Through pan-cancer analysis, we identified that GPRASP1 plays a critical role in the occurrence and prognosis of PC, and is closely related to immunological characteristics in PC. IHC analysis confirmed that GPRASP1 is significantly down-regulated in PC compared with normal tissues. The expression of GPRASP1 is significantly negatively correlated with clinical features (histologic grade, T stage, and TNM stage), and is an independent predictor of favorable prognosis, regardless of other clinicopathological features (HR: 0.69, 95% CI 0.54-0.92, p= 0.011). The etiological investigation found that the abnormal expression of GPRASP1 was related to DNA methylation and CNV frequency. Subsequently, the high expression of GPRASP1 was significantly correlated with immune cell infiltration (CD8+ T cell, tumor-infiltrating lymphocyte(TIL)), immune-related pathways(cytolytic activity, check-point, human leukocyte antigen (HLA)), immune checkpoint inhibitors (CTLA4, HAVCR2, LAG3, PDCD1 and TIGIT), immunomodulators ( CCR4/5/6, CXCL9, CXCR4/5), and immunogenicity(immune score, neoantigen, TMB(tumor mutation burden)). Finally, IPS (immunophenoscore) and TIDE (tumor immune dysfunction and exclusion) analysis demonstrated that GPRASP1 expression levels can accurately predict the immunotherapeutic response. GPRASP1 is a promising candidate biomarker that plays a role in the occurrence, development, and prognosis of PC. Evaluating GPRASP1 expression will aid in the characterization of tumor microenvironment (TME) infiltration and orient more efficient immunotherapy strategies.

Copy number variations in SPAST and ATL1 are rare among Brazilians.

Copy number variations (CNV) may represent a significant proportion of SPG4 and SPG3A diagnosis, the most frequent autosomal dominant subtypes of hereditary spastic paraplegias (HSP). We aimed to assess the frequency of CNVs in SPAST and ATL1 and to update the molecular epidemiology of HSP families in southern Brazil. A cohort study that included 95 Brazilian index cases with clinical suspicion of HSP was conducted between April 2011 and September 2022. Multiplex Ligation Dependent Probe Amplification (MLPA) was performed in 41 cases without defined diagnosis by different massive parallel sequencing techniques (MPS). Diagnosis was obtained in 57/95 (60%) index cases, 15/57 (26.3%) being SPG4. Most frequent autosomal recessive HSP subtypes were SPG7 followed by SPG11, SPG76 and cerebrotendinous xanthomatosis. No CNVs in SPAST and ATL1 were found. Copy number variations are rare among SPG4 and SPG3A families in Brazil. Considering the possibility of CNVs detection by specific algorithms with MPS data, we consider that this is likely the most cost-effective approach to investigate CNVs in these genes in low-risk populations, with MLPA being reserved as an orthogonal confirmatory test.