Matrisome Transcriptome Dynamics during Tissue Aging.

The extracellular matrix (ECM) is a complex three-dimensional network of macromolecules that provides structural support for the cells and plays a significant role in tissue homeostasis and repair. Growing evidence indicates that dysregulation of ECM remodeling contributes to various pathological conditions in the body, including age-associated diseases. In this work, gene expression data of normal human tissues obtained from the Genotype-Tissue Expression project, as well as data from MatrisomeDB 2.0, the ECM-protein knowledge database, are used to estimate the age-dependent matrisome transcriptome dynamics in the blood, heart, brain, liver, kidneys, lungs, and muscle. Differential gene expression (DE) analysis revealed dozens of matrisome genes encoding both structural elements of the ECM and ECM-associated proteins, which had a tissue-specific expression profile with age. Among common DE genes that changed expression with age in at least three tissues, COL18A1, MFAP1, IGFBP7, AEBP1, LTBP2, LTBP4, LG14, EFEMP1, PRELP, BGN, FAM20B, CTSC, CTSS, and CLEC2B were observed. The findings of the study also reveal that there are sex-specific alterations during aging in the matrisome gene expression. Taken together, the results obtained in this work may help in understanding the role of the ECM in tissue aging and might prove valuable for the future development of the field of ECM research in general.

Genetic changes in the FH gene cause vagal paraganglioma.

Vagal paraganglioma (VPGL) is a rare neuroendocrine tumor that originates from the paraganglion associated with the vagus nerve. VPGLs present challenges in terms of diagnostics and treatment. VPGL can occur as a hereditary tumor and, like other head and neck paragangliomas, is most frequently associated with mutations in the SDHx genes. However, data regarding the genetics of VPGL are limited. Herein, we report a rare case of a 41-year-old woman with VPGL carrying a germline variant in the FH gene. Using whole-exome sequencing, a variant, FH p.S249R, was identified; no variants were found in other PPGL susceptibility and candidate genes. Loss of heterozygosity analysis revealed the loss of the wild-type allele of the FH gene in the tumor. The pathogenic effect of the p.S249R variant on FH activity was confirmed by immunohistochemistry for S-(2-succino)cysteine (2SC). Potentially deleterious somatic variants were found in three genes, SLC7A7, ZNF225, and MED23. The latter two encode transcriptional regulators that can impact gene expression deregulation and are involved in tumor development and progression. Moreover, FH-mutated VPGL was characterized by a molecular phenotype different from SDHx-mutated PPGLs. In conclusion, the association of genetic changes in the FH gene with the development of VPGL was demonstrated. The germline variant FH: p.S249R and somatic deletion of the second allele can lead to biallelic gene damage that promotes tumor initiation. These results expand the clinical and mutation spectra of FH-related disorders and improve our understanding of the molecular genetic mechanisms underlying the pathogenesis of VPGL.

Decelerated Epigenetic Aging in Long Livers.

Epigenetic aging is a hot topic in the field of aging research. The present study estimated epigenetic age in long-lived individuals, who are currently actively being studied worldwide as an example of successful aging due to their longevity. We used Bekaert's blood-based age prediction model to estimate the epigenetic age of 50 conditionally "healthy" and 45 frail long-livers over 90 years old. Frailty assessment in long-livers was conducted using the Frailty Index. The control group was composed of 32 healthy individuals aged 20-60 years. The DNA methylation status of 4 CpG sites (ASPA CpG1, PDE4C CpG1, ELOVL2 CpG6, and EDARADD CpG1) included in the epigenetic clock was assessed through pyrosequencing. According to the model calculations, the epigenetic age of long-livers was significantly lower than their chronological age (on average by 21 years) compared with data from the group of people aged 20 to 60 years. This suggests a slowing of epigenetic and potentially biological aging in long livers. At the same time, the obtained results showed no statistically significant differences in delta age (difference between the predicted and chronological age) between "healthy" long livers and long livers with frailty. We also failed to detect sex differences in epigenetic age either in the group of long livers or in the control group. It is possible that the predictive power of epigenetic clocks based on a small number of CpG sites is insufficient to detect such differences. Nevertheless, this study underscores the need for further research on the epigenetic status of centenarians to gain a deeper understanding of the factors contributing to delayed aging in this population.

Transcriptome Profiling of Prostate Cancer, Considering Risk Groups and the TMPRSS2-ERG Molecular Subtype.

Molecular heterogeneity in prostate cancer (PCa) is one of the key reasons underlying the differing likelihoods of recurrence after surgical treatment in individual patients of the same clinical category. In this study, we performed RNA-Seq profiling of 58 localized PCa and 43 locally advanced PCa tissue samples obtained as a result of radical prostatectomy on a cohort of Russian patients. Based on bioinformatics analysis, we examined features of the transcriptome profiles within the high-risk group, including within the most commonly represented molecular subtype, TMPRSS2-ERG. The most significantly affected biological processes in the samples were also identified, so that they may be further studied in the search for new potential therapeutic targets for the categories of PCa under consideration. The highest predictive potential was found with the EEF1A1P5, RPLP0P6, ZNF483, CIBAR1, HECTD2, OGN, and CLIC4 genes. We also reviewed the main transcriptome changes in the groups at intermediate risk of PCa-Gleason Score 7 (groups 2 and 3 according to the ISUP classification)-on the basis of which the LPL, MYC, and TWIST1 genes were identified as promising additional prognostic markers, the statistical significance of which was confirmed using qPCR validation.

Transcriptomic Analysis of the Effect of Torin-2 on the Central Nervous System of Drosophila melanogaster.

Torin-2, a synthetic compound, is a highly selective inhibitor of both TORC1 and TORC2 (target of rapamycin) complexes as an alternative to the well-known immunosuppressor, geroprotector, and potential anti-cancer natural compound rapamycin. Torin-2 is effective at hundreds of times lower concentrations and prevents some negative side effects of rapamycin. Moreover, it inhibits the rapamycin-resistant TORC2 complex. In this work, we evaluated transcriptomic changes in D. melanogaster heads induced with lifetime diets containing Torin-2 and suggested possible neuroprotective mechanisms of Torin-2. The analysis included D. melanogaster of three ages (2, 4, and 6 weeks old), separately for males and females. Torin-2, taken at the lowest concentration being tested (0.5 µM per 1 L of nutrient paste), had a slight positive effect on the lifespan of D. melanogaster males (+4% on the average) and no positive effect on females. At the same time, RNA-Seq analysis revealed interesting and previously undiscussed effects of Torin-2, which differed between sexes as well as in flies of different ages. Among the cellular pathways mostly altered by Torin-2 at the gene expression level, we identified immune response, protein folding (heat shock proteins), histone modification, actin cytoskeleton organization, phototransduction and sexual behavior. Additionally, we revealed that Torin-2 predominantly reduced the expression of Srr gene responsible for the conversion of L-serine to D-serine and thus regulating activity of NMDA receptor. Via western blot analysis, we showed than in old males Torin-2 tends to increase the ratio of the active phosphorylated form of ERK, the lowest node of the MAPK cascade, which may play a significant role in neuroprotection. Thus, the complex effect of Torin-2 may be due to the interplay of the immune system, hormonal background, and metabolism. Our work is of interest for further research in the field of NMDA-mediated neurodegeneration.

Lymphatic Dissemination in Prostate Cancer: Features of the Transcriptomic Profile and Prognostic Models.

Radical prostatectomy is the gold standard treatment for prostate cancer (PCa); however, it does not always completely cure PCa, and patients often experience a recurrence of the disease. In addition, the clinical and pathological parameters used to assess the prognosis and choose further tactics for treating a patient are insufficiently informative and need to be supplemented with new markers. In this study, we performed RNA-Seq of PCa tissue samples, aimed at identifying potential prognostic markers at the level of gene expression and miRNAs associated with one of the key signs of cancer aggressiveness-lymphatic dissemination. The relative expression of candidate markers was validated by quantitative PCR, including an independent sample of patients based on archival material. Statistically significant results, derived from an independent set of samples, were confirmed for miR-148a-3p and miR-615-3p, as well as for the CST2, OCLN, and PCAT4 genes. Considering the obtained validation data, we also analyzed the predictive value of models based on various combinations of identified markers using algorithms based on machine learning. The highest predictive potential was shown for the "CST2 + OCLN + pT" model (AUC = 0.863) based on the CatBoost Classifier algorithm.

ALDH3A2, ODF2, QSOX2, and MicroRNA-503-5p Expression to Forecast Recurrence in TMPRSS2-ERG-Positive Prostate Cancer.

Following radical surgery, patients may suffer a relapse. It is important to identify such patients so that therapy tactics can be modified appropriately. Existing stratification schemes do not display the probability of recurrence with enough precision since locally advanced prostate cancer (PCa) is classified as high-risk but is not ranked in greater detail. Between 40 and 50% of PCa cases belong to the TMPRSS2-ERG subtype that is a sufficiently homogeneous group for high-precision prognostic marker search to be possible. This study includes two independent cohorts and is based on high throughput sequencing and qPCR data. As a result, we have been able to suggest a perspective-trained model involving a deep neural network based on both qPCR data for mRNA and miRNA and clinicopathological criteria that can be used for recurrence risk forecasts in patients with TMPRSS2-ERG-positive, locally advanced PCa (the model uses ALDH3A2 + ODF2 + QSOX2 + hsa-miR-503-5p + ISUP + pT, with an AUC = 0.944). In addition to the prognostic model's use of identified differentially expressed genes and miRNAs, miRNA-target pairs were found that correlate with the prognosis and can be presented as an interactome network.

Dynamic Profiling of Exosomal microRNAs in Blood Plasma of Patients with Castration-Resistant Prostate Cancer.

Prostate cancer is one of the most common and socially significant cancers among men. The aim of this study was to identify significant changes in the expression of exosomal miRNAs associated with an increase in the level of prostate specific antigen in castration-resistant prostate cancer during therapy and to evaluate them as potential prognostic markers for this category of disease. High-throughput miRNA sequencing was performed on 49 blood plasma samples taken from 11 Russian patients with castration-resistant cancer during therapy. Bioinformatic analysis of the obtained miRNA-seq data was carried out. Additionally, miRNA-seq data from the PRJNA562276 project were analyzed to identify exosomal miRNAs associated with castration-resistant prostate cancer. We found 34 differentially expressed miRNAs associated with the progression of castration-resistant prostate cancer during therapy in Russian patients. It was also shown that hsa-miRNA-148a-3p expression can serve as a potential prognostic marker. We found the exosomal miRNA expression signature associated with castration-resistant prostate cancer progression, in particular on the Russian patient cohort. Many of these miRNAs are well-known players in either oncogenic transformation or tumor suppression. Further experimental studies with extended sampling are required to validate these results.

Impact TMPRSS2-ERG Molecular Subtype on Prostate Cancer Recurrence.

Currently, seven molecular subtypes of prostate cancer (PCa) are known, the most common of which being the subtype characterized by the presence of the TMPRSS2-ERG fusion transcript. While there is a considerable amount of work devoted to the influence of this transcript on the prognosis of the disease, data on its role in the progression and prognosis of PCa remain controversial. The present study is devoted to the analysis of the association between the TMPRSS2-ERG transcript and the biochemical recurrence of PCa. The study included two cohorts: the RNA-Seq sample of Russian patients with PCa (n = 72) and the TCGA-PRAD data (n = 203). The results of the analysis of the association between the TMPRSS2-ERG transcript and biochemical recurrence were contradictory. The differential expression analysis (biochemical recurrence cases versus biochemical recurrence-free) and the gene set enrichment analysis revealed a list of genes involved in major cellular pathways. The GNL3, QSOX2, SSPO, and SYS1 genes were selected as predictors of the potential prognostic model (AUC = 1.000 for a cohort of Russian patients with PCa and AUC = 0.779 for a TCGA-PRAD cohort).

De Novo Transcriptome Profiling of Brain Tissue from the Annual Killifish Nothobranchius guentheri.

Nothobranchius is a genus of small annual killifish found in Africa. Due to the relatively short lifespan, as well as easy breeding and care, Nothobranchius fish are becoming widely used as a vertebrate model system. Studying the genome and transcriptome of these fish is essential for advancing the field. In this study, we performed de novo transcriptome assembly of brain tissues from Nothobranchius guentheri using Trinity. Annotation of 104,271 potential genes (with transcripts longer than 500 bp) was carried out; for 24,967 genes (53,654 transcripts), in which at least one GO annotation was derived. We also analyzed the effect of a long-term food supplement with Torin 2, second-generation ATP-competitive inhibitor of mTOR, on the gene expression changes in brain tissue of adult N. guentheri. Overall, 1491 genes in females and 249 genes in males were differently expressed under Torin 2-supplemented diet. According to the Gene Set Enrichment Analysis (GSEA), the majority of identified genes were predominantly involved in the regulation of metabolic process, dendritic spine maintenance, circadian rhythms, retrotransposition, and immune response. Thus, we have provided the first transcriptome assembly and assessed the differential gene expression in response to exposure to Torin 2, which allow a better understanding of molecular changes in the brain tissues of adult fish in the mTOR pathway inhibition.

NETO2 Is Deregulated in Breast, Prostate, and Colorectal Cancer and Participates in Cellular Signaling.

The NETO2 gene (neuropilin and tolloid-like 2) encodes a protein that acts as an accessory subunit of kainate receptors and is predominantly expressed in the brain. Upregulation of NETO2 has been observed in several tumors; however, its role in tumorigenesis remains unclear. In this study, we investigated NETO2 expression in breast, prostate, and colorectal cancer using quantitative PCR (qPCR), as well as the effect of shRNA-mediated NETO2 silencing on transcriptome changes in colorectal cancer cells. In the investigated tumors, we observed both increased and decreased NETO2 mRNA levels, presenting no correlation with the main clinicopathological characteristics. In HCT116 cells, NETO2 knockdown resulted in the differential expression of 17 genes and 2 long non-coding RNAs (lncRNAs), associated with the upregulation of circadian rhythm and downregulation of several cancer-associated pathways, including Wnt, transforming growth factor (TGF)-ß, Janus kinase (JAK)-signal transducer and activator of transcription (STAT), mitogen-activated protein kinase (MAPK), and phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) pathways. Furthermore, we demonstrated the possibility to utilize a novel model organism, short-lived fish Nothobranchius furzeri, for evaluating NETO2 functions. The ortholog neto2b in N. furzeri demonstrated a high similarity in nucleotide and amino acid sequences with human NETO2, as well as was characterized by stable expression in various fish tissues. Collectively, our findings demonstrate the deregulation of NETO2 in the breast, prostate, and colorectal cancer and its participation in the tumor development primarily through cellular signaling.

Multiple paragangliomas: a case report.

BACKGROUND: Carotid and vagal paragangliomas (CPGLs and VPGLs) are rare neoplasms that arise from the paraganglia located at the bifurcation of carotid arteries and vagal trunk, respectively. Both tumors can occur jointly as multiple paragangliomas accounting for approximately 10 to 20% of all head and neck paragangliomas. However, molecular and genetic mechanisms underlying the pathogenesis of multiple paragangliomas remain elusive. CASE PRESENTATION: We report a case of multiple paragangliomas in a patient, manifesting as bilateral CPGL and unilateral VPGL. Tumors were revealed via computed tomography and ultrasound study and were resected in two subsequent surgeries. Both CPGLs and VPGL were subjected to immunostaining for succinate dehydrogenase (SDH) subunits and exome analysis. A likely pathogenic germline variant in the SDHD gene was indicated, while likely pathogenic somatic variants differed among the tumors. CONCLUSIONS: The identified germline variant in the SDHD gene seems to be a driver in the development of multiple paragangliomas. However, different spectra of somatic variants identified in each tumor indicate individual molecular mechanisms underlying their pathogenesis.

Mutation profiling in eight cases of vagal paragangliomas.

BACKGROUND: Vagal paragangliomas (VPGLs) belong to a group of rare head and neck neuroendocrine tumors. VPGLs arise from the vagus nerve and are less common than carotid paragangliomas. Both diagnostics and therapy of the tumors raise significant challenges. Besides, the genetic and molecular mechanisms behind VPGL pathogenesis are poorly understood. METHODS: The collection of VPGLs obtained from 8 patients of Russian population was used in the study. Exome library preparation and high-throughput sequencing of VPGLs were performed using an Illumina technology. RESULTS: Based on exome analysis, we identified pathogenic/likely pathogenic variants of the SDHx genes, frequently mutated in paragangliomas/pheochromocytomas. SDHB variants were found in three patients, whereas SDHD was mutated in two cases. Moreover, likely pathogenic missense variants were also detected in SDHAF3 and SDHAF4 genes encoding for assembly factors for the succinate dehydrogenase (SDH) complex. In a patient, we found a novel variant of the IDH2 gene that was predicted as pathogenic by a series of algorithms used (such as SIFT, PolyPhen2, FATHMM, MutationTaster, and LRT). Additionally, pathogenic/likely pathogenic variants were determined for several genes, including novel genes and some genes previously reported as associated with different types of tumors. CONCLUSIONS: Results indicate a high heterogeneity among VPGLs, however, it seems that driver events in most cases are associated with mutations in the SDHx genes and SDH assembly factor-coding genes that lead to disruptions in the SDH complex.

miRNAs expression signature potentially associated with lymphatic dissemination in locally advanced prostate cancer.

BACKGROUND: Prostate cancer is one of the most common and socially significant cancers among men. The aim of our study was to reveal changes in miRNA expression profiles associated with lymphatic dissemination in prostate cancer and to identify the most prominent miRNAs as potential prognostic markers for future studies. METHODS: High-throughput miRNA sequencing was performed for 44 prostate cancer specimens taken from Russian patients, with and without lymphatic dissemination (N1 - 20 samples; N0 - 24 samples). RESULTS: We found at least 18 microRNAs with differential expression between N0 and N1 sample groups: miR-182-5p, miR-183-5p, miR-96-5p, miR-25-3p, miR-93-5p, miR-7-5p, miR-615-3p, miR-10b, miR-1248 (N1-miRs; elevated expression in N1 cohort; p < 0.05); miR-1271-5p, miR-184, miR-222-3p, miR-221-5p, miR-221-3p, miR-455-3p, miR-143-5p, miR-181c-3p and miR-455-5p (N0-miRs; elevated expression in N0; p < 0.05). The expression levels of N1-miRs were highly correlated between each other (the same is applied for N0-miRs) and the expression levels of N0-miRs and N1-miRs were anti-correlated. The tumor samples can be divided into two groups depending on the expression ratio between N0-miRs and N1-miRs. CONCLUSIONS: We found the miRNA expression signature associated with lymphatic dissemination, in particular on the Russian patient cohort. Many of these miRNAs are well-known players in either oncogenic transformation or tumor suppression. Further experimental studies with extended sampling are required to validate these results.

Immunohistochemistry and Mutation Analysis of SDHx Genes in Carotid Paragangliomas.

Carotid paragangliomas (CPGLs) are rare neuroendocrine tumors often associated with mutations in SDHx genes. The immunohistochemistry of succinate dehydrogenase (SDH) subunits has been considered a useful instrument for the prediction of SDHx mutations in paragangliomas/pheochromocytomas. We compared the mutation status of SDHx genes with the immunohistochemical (IHC) staining of SDH subunits in CPGLs. To identify pathogenic/likely pathogenic variants in SDHx genes, exome sequencing data analysis among 42 CPGL patients was performed. IHC staining of SDH subunits was carried out for all CPGLs studied. We encountered SDHx variants in 38% (16/42) of the cases in SDHx genes. IHC showed negative (5/15) or weak diffuse (10/15) SDHB staining in most tumors with variants in any of SDHx (94%, 15/16). In SDHA-mutated CPGL, SDHA expression was completely absent and weak diffuse SDHB staining was detected. Positive immunoreactivity for all SDH subunits was found in one case with a variant in SDHD. Notably, CPGL samples without variants in SDHx also demonstrated negative (2/11) or weak diffuse (9/11) SDHB staining (42%, 11/26). Obtained results indicate that SDH immunohistochemistry does not fully reflect the presence of mutations in the genes; diagnostic effectiveness of this method was 71%. However, given the high sensitivity of SDHB immunohistochemistry, it could be used for initial identifications of patients potentially carrying SDHx mutations for recommendation of genetic testing.

Gene Expression Changes and Associated Pathways Involved in the Progression of Prostate Cancer Advanced Stages.

Prostate cancer (PC) is one of the most common cancers among men worldwide, and advanced PCs, such as locally advanced PC (LAPC) and castration-resistant PC (CRPC), present the greatest challenges in clinical management. Current indicators have limited capacity to predict the disease course; therefore, better prognostic markers are greatly needed. In this study, we performed a bioinformatic analysis of The Cancer Genome Atlas (TCGA) datasets, including RNA-Seq data from the prostate adenocarcinoma (PRAD; n = 55) and West Coast Dream Team - metastatic CRPC (WCDT-MCRPC; n = 84) projects, to evaluate the transcriptome changes associated with progression-free survival (PFS) for LAPC and CRPC, respectively. We identified the genes whose expression was positively/negatively correlated with PFS. In LAPC, the genes with the most significant negative correlations were ZC2HC1A, SQLE, and KIF11, and the genes with the most significant positive correlations were SOD3, LRRC26, MIR22HG, MEG3, and MIR29B2CHG. In CRPC, the most significant positive correlations were found for BET1, CTAGE5, IFNGR1, and GIMAP6, and the most significant negative correlations were found for CLPB, PRPF19, ZNF610, MPST, and LINC02001. In addition, we performed a gene network interaction analysis using STRINGdb, which revealed a significant relationship between genes predominantly involved in the cell cycle and characterized by upregulated expression in early recurrence. Based on the results, we propose several genes that can be used as potential prognostic markers.

Mutation Frequency in Main Susceptibility Genes Among Patients With Head and Neck Paragangliomas.

Head and neck paragangliomas (HNPGLs) are rare neuroendocrine tumors that have a high degree of heritability and are predominantly associated with mutations in ten genes, such as SDHx, SDHAF2, VHL, RET, NF1, TMEM127, MAX, FH, MEN2, and SLC25A11. Elucidating the mutation prevalence is crucial for the development of genetic testing. In this study, we identified pathogenic/likely pathogenic variants in the main susceptibility genes in 102 Russian patients with HNPGLs (82 carotid and 23 vagal paragangliomas) using whole exome sequencing. Pathogenic/likely pathogenic variants were detected in 43% (44/102) of patients. We identified the following variant distribution of the tested genes: SDHA (1%), SDHB (10%), SDHC (5%), SDHD (24.5%), and RET (5%). SDHD variants were observed in the majority of the patients with bilateral/multiple paragangliomas. Thus, among Russian patients with HNPGLs the most frequently mutated gene was SDHD followed by SDHB, SDHC, RET, and SDHA.

Differentially Expressed Genes Associated With Prognosis in Locally Advanced Lymph Node-Negative Prostate Cancer.

Older age is one of the main risk factors for cancer development. The incidence of prostate cancer, as a multifactorial disease, also depends upon demographic factors, race, and genetic predisposition. Prostate cancer most frequently occurs in men over 60 years of age, indicating a clear association between older age and disease onset. Carcinogenesis is followed by the deregulation of many genes, and some of these changes could serve as biomarkers for diagnosis, prognosis, prediction of drug therapy efficacy, as well as possible therapeutic targets. We have performed a bioinformatic analysis of a The Cancer Genome Atlas (TCGA) data and RNA-Seq profiling of a Russian patient cohort to reveal prognostic markers of locally advanced lymph node-negative prostate cancer (lymph node-negative LAPC). We also aimed to identify markers of the most common molecular subtype of prostate cancer carrying a fusion transcript TMPRSS2-ERG. We have found several genes that were differently expressed between the favorable and unfavorable prognosis groups and involved in the enriched KEGG pathways based on the TCGA (B4GALNT4, PTK6, and CHAT) and Russian patient cohort data (AKR1C1 and AKR1C3). Additionally, we revealed such genes for the TMPRSS2-ERG prostate cancer molecular subtype (B4GALNT4, ASRGL1, MYBPC1, RGS11, SLC6A14, GALNT13, and ST6GALNAC1). Obtained results contribute to a better understanding of the molecular mechanisms behind prostate cancer progression and could be used for further development of the LAPC prognosis marker panel.

Novel potential causative genes in carotid paragangliomas.

BACKGROUND: Carotid paragangliomas (CPGLs) are rare neuroendocrine tumors that arise from the paraganglion at the bifurcation of the carotid artery and are responsible for approximately 65% of all head and neck paragangliomas. CPGLs can occur sporadically or along with different hereditary tumor syndromes. Approximately 30 genes are known to be associated with CPGLs. However, the genetic basis behind the development of these tumors is not fully elucidated, and the molecular mechanisms underlying CPGL pathogenesis remain unclear. METHODS: Whole exome and transcriptome high-throughput sequencing of CPGLs was performed on an Illumina platform. Exome libraries were prepared using a Nextera Rapid Capture Exome Kit (Illumina) and were sequenced under 75 bp paired-end model. For cDNA library preparation, a TruSeq Stranded Total RNA Library Prep Kit with Ribo-Zero Gold (Illumina) was used; transcriptome sequencing was carried out with 100 bp paired-end read length. Obtained data were analyzed using xseq which estimates the influence of mutations on gene expression profiles allowing to identify potential causative genes. RESULTS: We identified a total of 16 candidate genes (MYH15, CSP1, MYH3, PTGES3L, CSGALNACT2, NMD3, IFI44, GMCL1, LSP1, PPFIBP2, RBL2, MAGED1, CNIH3, STRA6, SLC6A13, and ATM) whose variants potentially influence their expression (cis-effect). The strongest cis-effect of loss-of-function variants was found in MYH15, CSP1, and MYH3, and several likely pathogenic variants in these genes associated with CPGLs were predicted. CONCLUSIONS: Using the xseq probabilistic model, three novel potential causative genes, namely MYH15, CSP1, and MYH3, were identified in carotid paragangliomas.

The CIMP-high phenotype is associated with energy metabolism alterations in colon adenocarcinoma.

BACKGROUND: CpG island methylator phenotype (CIMP) is found in 15-20% of malignant colorectal tumors and is characterized by strong CpG hypermethylation over the genome. The molecular mechanisms of this phenomenon are not still fully understood. The development of CIMP is followed by global gene expression alterations and metabolic changes. In particular, CIMP-low colon adenocarcinoma (COAD), predominantly corresponded to consensus molecular subtype 3 (CMS3, "Metabolic") subgroup according to COAD molecular classification, is associated with elevated expression of genes participating in metabolic pathways. METHODS: We performed bioinformatics analysis of RNA-Seq data from The Cancer Genome Atlas (TCGA) project for CIMP-high and non-CIMP COAD samples with DESeq2, clusterProfiler, and topGO R packages. Obtained results were validated on a set of fourteen COAD samples with matched morphologically normal tissues using quantitative PCR (qPCR). RESULTS: Upregulation of multiple genes involved in glycolysis and related processes (ENO2, PFKP, HK3, PKM, ENO1, HK2, PGAM1, GAPDH, ALDOA, GPI, TPI1, and HK1) was revealed in CIMP-high tumors compared to non-CIMP ones. Most remarkably, the expression of the PKLR gene, encoding for pyruvate kinase participating in gluconeogenesis, was decreased approximately 20-fold. Up to 8-fold decrease in the expression of OGDHL gene involved in tricarboxylic acid (TCA) cycle was observed in CIMP-high tumors. Using qPCR, we confirmed the increase (4-fold) in the ENO2 expression and decrease (2-fold) in the OGDHL mRNA level on a set of COAD samples. CONCLUSIONS: We demonstrated the association between CIMP-high status and the energy metabolism changes at the transcriptomic level in colorectal adenocarcinoma against the background of immune pathway activation. Differential methylation of at least nine CpG sites in OGDHL promoter region as well as decreased OGDHL mRNA level can potentially serve as an additional biomarker of the CIMP-high status in COAD.