153 research outputs found
Gene length corrected trimmed mean of M-values (GeTMM) processing of RNA-seq data performs similarly in intersample analyses while improving intrasample comparisons
Background: Current normalization methods for RNA-sequencing data allow either for intersample comparison to identify differentially expressed (DE) genes or for intrasample comparison for the discovery and validation of gene signatures. Most studies on optimization of normalization methods typically use simulated data to validate methodologies. We describe a new method, GeTMM, which allows for both inter- and intrasample analyses with the same normalized data set. We used actual (i.e. not simulated) RNA-seq data from 263 colon cancers (no biological replicates) and used the same read count data to compare GeTMM with the most commonly used normalization methods (i.e. TMM (used by edgeR), RLE (used by DESeq2) and TPM) with respect to distributions, effect of RNA quality, subtype-classification, recurrence score, recall of DE genes and correlation to RT-qPCR data. Results: We observed a clear benefit for GeTMM and TPM with regard to intrasample comparison while GeTMM performed similar to TMM and RLE normalized data in intersample comparisons. Regarding DE genes, recall was found comparable among the normalization methods, while GeTMM showed the lowest number of false-positive DE genes. Remarkably, we observed limited detrimental effects in samples with low RNA quality. Conclusions: We show that GeTMM outperforms established methods with regard to intrasample comparison while performing equivalent with regard to intersample normalization using the same normalized data. These combined properties enhance the general usefulness of RNA-seq but also the comparability to the many array-based gene expression data in the public domain
mRNA expression profiles in circulating tumor cells of metastatic colorectal cancer patients
The molecular characterization of circulating tumor cells (CTCs) is a promising tool for the repeated and non-invasive evaluation of predictive and prognostic factors. Challenges associated with CTC characterization using the only FDA approved method for CTC enumeration, the CellSearch technique, include the presence of an excess of leukocytes in CTC-enriched blood fractions. Here we aimed to identify colorectal tumor-specific gene expression levels in the blood of patients with and without detectable CTCs according to CellSearch criteria. Materials and methods: Blood of 30 healthy donors (HDs) and 142 metastatic colorectal cancer (mCRC) patients was subjected to CellSearch CTC enumeration and isolation. In all samples, 95 mRNAs were measured by reverse transcriptase quantitative PCR (RT-qPCR). HD blood samples and patient samples with three or more CTCs were compared to identify CTC-specific mRNAs. Patient samples without detectable CTCs were separately analyzed. Results: Thirty-four CTC-specific mRNAs were higher expressed in patients with â„3 CTCs compared with HDs (Mann-Whitney U-test P<0.05). Among patients without detectable CTCs, a HD-unlike subgroup was identified which could be distinguished from HDs by the expression of epithelial genes such as KRT19, KRT20 and AGR2. Also, in an independent patient set, a similar HD-unlike group could be identified among the patients without detectable CTCs according to the CellSearch system. Conclusion: Extensive molecular characterization of colorectal CTCs is feasible and a subgroup of patients without detectable CTCs according to CellSearch criteria bears circulating tumor load, which may have clinical consequences. This CTC-specific gene panel for mCRC patients may enable the exploration of CTC characterization as a novel means to further individualize cancer treatment
Whole Gene Capture Analysis of 15 CRC Susceptibility Genes in Suspected Lynch Syndrome Patients
Background and Aims Lynch Syndrome (LS) is caused by pathogenic germline variants in one of the mismatch repair (MMR) genes. However, up to 60% of MMR-deficient colorectal cancer cases are categorized as suspected Lynch Syndrome (sLS) because no pathogenic MMR germline variant can be identified, which leads to difficulties in clinical management. We therefore analyzed the genomic regions of 15 CRC susceptibility genes in leukocyte DNA of 34 unrelated sLS patients and 11 patients with MLH1 hypermethylated tumors with a clear family history. Methods Using targeted next-generation sequencing, we analyzed the entire non-repetitive genomic sequence, including intronic and regulatory sequences, of 15 CRC susceptibility genes. In addition, tumor DNA from 28 sLS patients was analyzed for somatic MMR variants. Results Of 1979 germline variants found in the leukocyte DNA of 34 sLS patients, one was a pathogenic variant (MLH1 c.1667+1delG). Leukocyte DNA of 11 patients with MLH1 hypermethylated tumors was negative for pathogenic germline variants in the tested CRC susceptibility genes and for germline MLH1 hypermethylation. Somatic DNA analysis of 28 sLS tumors identified eight (29%) cases with two pathogenic somatic variants, one with a VUS predicted to pathogenic and LOH, and nine cases (32%) with one pathogenic somatic variant (n = 8) or one VUS predicted to be pathogenic (n = 1). Conclusions This is the first study in sLS patients to include the entire genomic sequence of CRC susceptibility genes. An underlying somatic or germline MMR gene defect was identified in ten of 34 sLS patients (29%). In the remaining sLS patients, the underlying genetic defect explaining the MMRdeficiency in their tumors might be found outside the genomic regions harboring the MMR and other known CRC susceptibility genes
Confirmation of a metastasis-specific microRNA signature in primary colon cancer
The identification of patients with high-risk stage II colon cancer who may benefit from adjuvant therapy may allow the clinical approach to be tailored for these patients based on an understanding of tumour biology. MicroRNAs have been proposed as markers of the prognosis or treatment response in colorectal cancer. Recently, a 2-microRNA signature (l et-7i and miR-10b) was proposed to identify colorectal cancer patients at risk of developing distant metastasis. We assessed the prognostic value of this signature and additional candidate microRNAs in an independent, clinically well-defined, prospectively collected cohort of primary colon cancer patients including stage I-II colon cancer without and stage III colon cancer with adjuvant treatment. The 2-microRNA signature specifically predicted hepatic recurrence in the stage I-II group, but not the overall ability to develop distant metastasis. The addition of miR-30b to the 2-microRNA signature allowed the prediction of both distant metastasis and hepatic recurrence in patients with stage I-II colon cancer who did not receive adjuvant chemotherapy. Available gene expression data allowed us to associate m iR-30b expression with axon guidance and l et-7i expression with cell adhesion, migration, and motility
Molecular characteristics of circulating tumor cells resemble the liver metastasis more closely than the primary tumor in metastatic colorectal cancer
Background: CTCs are a promising alternative for metastatic tissue biopsies for use in precision medicine approaches. We investigated to what extent the molecular characteristics of circula
An 8-gene mRNA expression profile in circulating tumor cells predicts response to aromatase inhibitors in metastatic breast cancer patients
Background: Molecular characterization of circulating tumor cells (CTC) is promising for personalized medicine. We aimed to identify a CTC gene expression profile predicting outcome to first-line aromatase inhibitors in metastatic breast cancer (MBC) patients. Methods: CTCs were isolated from 78 MBC patients before treatment start. mRNA expression levels of 96 genes were measured by quantitative reverse transcriptase polymerase chain reaction. After applying predefined exclusion criteria based on lack of sufficient RNA quality and/or quantity, the data from 45 patients were used to construct a gene expression profile to predict poor responding patients, defined as disease progression or death <9 months, by a leave-one-out cross validation. Results: Of the 45 patients, 19 were clinically classified as poor responders. To identify them, the 75 % most variable genes were used to select genes differentially expressed between good and poor responders. An 8-gene CTC predictor was significantly associated with outcome (Hazard Ratio [HR] 4.40, 95 % Confidence Interval [CI]: 2.17-8.92, P < 0.001). This predictor identified poor responding patients with a sensitivity of 63 % and a positive predictive value of 75 %, while good responding patients were correctly predicted in 85 % of the cases. In multivariate Cox regression analysis, including CTC count at baseline, the 8-gene CTC predictor was the only factor independently associated with outcome (HR 4.59 [95 % CI: 2.11-9.56], P < 0.001). This 8-gene signature was not associated with outcome in a group of 71 MBC patients treated with systemic treatments other than AI. Conclusions: An 8-gene CTC predictor was identified which discriminates good and poor outcome to first-line aromatase inhibitors in MBC patients. Although results need to be validated, this study underscores the potential of molecular characterization of CTCs
Clinical significance of the nuclear receptor co-regulator DC-SCRIPT in breast cancer: an independent retrospective validation study
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Igbo-English intrasentential codeswitching and the Matrix Language Frame model
This paper uses data from Igbo-English intrasentential codeswitching involving mixed nominal expressions to test the Matrix Language Frame (MLF) model. The MLF model is one of the most highly influential frameworks used in the study of grammatical aspects of codeswitching. Three principles associated with it, the Matrix Language Principle, the Asymmetry Principle and the Uniform Structure Principle, were tested on data collected from informal conversations by educated adult Igbo-English bilinguals resident in Port Harcourt. The results of the analyses suggest general support for the three principles and for identifying Igbo-English as a âclassicâ case of codeswitching
Diversity in African languages: Selected papers from the 46th Annual Conference on African Linguistics
Diversity in African Languages contains a selection of revised papers from the 46th Annual Conference on African Linguistics, held at the University of Oregon. Most chapters focus on single languages, addressing diverse aspects of their phonology, morphology, semantics, syntax, information structure, or historical development. These chapters represent nine different genera: Mande, Gur, Kwa, Edoid, Bantu, Nilotic, Gumuzic, Cushitic, and Omotic. Other chapters investigate a mix of languages and families, moving from typological issues to sociolinguistic and inter-ethnic factors that affect language and accent switching. Some chapters are primarily descriptive, while others push forward the theoretical understanding of tone, semantic problems, discourse related structures, and other linguistic systems. The papers on Bantu languages reflect something of the internal richness and continued fascination of the family for linguists, as well as maturation of research on the family. The distribution of other papers highlights the need for intensified research into all the language families of Africa, including basic documentation, in order to comprehend linguistic diversities and convergences across the continent. In this regard, the chapter on DaatsâiÌin (Gumuzic) stands out as the first-ever published article on this hitherto unknown and endangered language found in the Ethiopian-Sudanese border lands
Diversity in African languages: Selected papers from the 46th Annual Conference on African Linguistics
Diversity in African Languages contains a selection of revised papers from the 46th Annual Conference on African Linguistics, held at the University of Oregon. Most chapters focus on single languages, addressing diverse aspects of their phonology, morphology, semantics, syntax, information structure, or historical development. These chapters represent nine different genera: Mande, Gur, Kwa, Edoid, Bantu, Nilotic, Gumuzic, Cushitic, and Omotic. Other chapters investigate a mix of languages and families, moving from typological issues to sociolinguistic and inter-ethnic factors that affect language and accent switching. Some chapters are primarily descriptive, while others push forward the theoretical understanding of tone, semantic problems, discourse related structures, and other linguistic systems. The papers on Bantu languages reflect something of the internal richness and continued fascination of the family for linguists, as well as maturation of research on the family. The distribution of other papers highlights the need for intensified research into all the language families of Africa, including basic documentation, in order to comprehend linguistic diversities and convergences across the continent. In this regard, the chapter on DaatsâiÌin (Gumuzic) stands out as the first-ever published article on this hitherto unknown and endangered language found in the Ethiopian-Sudanese border lands
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