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Different next-generation sequencing pipelines based detection of tumor DNA in cerebrospinal fluid of lung adenocarcinoma cancer patients with leptomeningeal metastases.

ABSTRACT: BACKGROUND:The nucleic acid mutation status in intracranial metastasis is markedly significant clinically. The goal of the current study was to explore whether the tumor-associated mutations can be detected by different next-generation sequencing (NGS) pipelines in paired cerebrospinal fluid (CSF) and plasma samples from lung adenocarcinoma (LAC) patients with leptomeningeal metastases (LM). METHODS:Paired CSF cell free DNA (cfDNA), CSF cells, plasma and formalin-fixed and paraffin-embedded (FFPE) samples of primary tumors were collected from 29 LAC patients with LM to detect the mutations by different NGS pipelines. RESULTS:DNA libraries were generated successfully for 79 various samples in total for NGS sequencing, of which mutations were detected in 7 plasma samples (24.14%), 12 CSF cfDNA samples (66.67%), and 10 CSF cells (76.9%) samples. For the 26 patients with detected mutations, 8/26(30.77%) had mutations in plasma, which was significantly lower than that those from CSF cfDNA (12/15, 80.00%), CSF cells (10/11, 90.91%) and FFPE samples (13/17, 76.47%). When the input DNA of CSF cells was less than 20?ng, the cHOPE pipeline of NGS identified the most mutations for epidermal growth factor receptor (EGFR). CONCLUSIONS:NGS-based detection of mutations in cfDNA or cells from CSF provided more information than from plasma samples from LAC patients with LM. In addition, the cHOPE pipeline performed better than the other three NGS pipelines when input DNA from CSF cells was low.

SUBMITTER: Ge M

PROVIDER: S-EPMC6373107 | biostudies-literature | 2019 Feb

REPOSITORIES: biostudies-literature

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Different next-generation sequencing pipelines based detection of tumor DNA in cerebrospinal fluid of lung adenocarcinoma cancer patients with leptomeningeal metastases.

Ge Mengxi M Zhan Qiong Q Zhang Zhenzhen Z Ji Xiaoyu X Zhou Xinli X Huang Ruofan R Liang Xiaohua X

BMC cancer 20190212 1

<h4>Background</h4>The nucleic acid mutation status in intracranial metastasis is markedly significant clinically. The goal of the current study was to explore whether the tumor-associated mutations can be detected by different next-generation sequencing (NGS) pipelines in paired cerebrospinal fluid (CSF) and plasma samples from lung adenocarcinoma (LAC) patients with leptomeningeal metastases (LM).<h4>Methods</h4>Paired CSF cell free DNA (cfDNA), CSF cells, plasma and formalin-fixed and paraffi ...[more]

PMID: 30755180

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Project description:Background: Leptomeningeal metastasis (LM) has frequently been observed in patients with lung adenocarcinoma. So far, its diagnosis and disease course monitoring are still extremely difficult. Moreover, there is no effective treatment regimen for LM due to a lack knowledge on the molecular mechanism of LM. This study aimed to identify LM-related cerebrospinal fluid (CSF) miRNAs, which have potential value for diagnosing and monitoring LM and exploring the molecular mechanism. Methods: CSF miRNAs were screened and verified by microarray analysis and quantitative real-time PCR (qRT-PCR) in LM patients with lung adenocarcinoma and non-LM controls, and the diagnostic performance of candidate miRNAs was evaluated. Then, candidate miRNAs in matched CSF samples from LM patients at diagnosis, after initial therapy, at relapse, and after salvage therapy, were analyzed to assess the relationship between CSF miRNAs and LM disease course. The effect of candidate miRNAs on proliferation, invasion, and migration of lung adenocarcinoma cell lines was assessed. The targeted genes of the candidate miRNA were predicted by TargetScan, miRDB, and miRTarbase online analysis tools. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) were used to analyze the functional categories of predicted target genes. Results: CSF miR-7975, miR-7977, and miR-7641 were screened and verified to be statistically significantly up-regulated in LM patients compared to non-LM controls. The three miRNAs, when combined, exhibited optimal diagnostic performance. Longitudinal data of CSF miR-7975 and miR-7977 correlated well with clinical courses of LM. Overexpression of miR-7977 promoted proliferation, migration, and invasion of lung adenocarcinoma cells. Moreover, 385 targeted genes of miR-7977 were predicted and were involved in various pathways related to cancer metastasis. Conclusions: This study offers insights for future research of CSF miRNAs as robust tools for diagnosing and monitoring LM. It also reveals a novel pathway for exploration of underlying mechanisms of LM.

Project description:PurposeThe application of metagenomic next-generation sequencing (mNGS) in the diagnosis of tuberculous meningitis (TBM) remains poorly characterized. Here, we retrospectively analyzed data from patients with TBM who had taken both mNGS and conventional tests including culture of Mycobacterium tuberculosis (MTB), polymerase chain reaction (PCR) and acid-fast bacillus (AFB) stain, and the sensitivity and specificity of these methods were compared.MethodsWe retrospectively recruited TBM patients admitted to the hospital between December 2015 and October 2018. The first collection of cerebrospinal fluid (CSF) samples underwent both mNGS and conventional tests. In addition, patients with bacterial/cryptococcal meningitis or viral meningoencephalitis were mNGS positive controls, and a patient with auto-immune encephalitis was an mNGS negative control.ResultsTwenty three TBM patients were classified as 12 definite and 11 clinical diagnoses, which were based on clinical manifestations, pathogen evidence, CSF parameters, brain imaging, and treatment response. The mNGS method identified sequences of Mycobacterium tuberculosis complex (MBTC) from 18 samples (18/23, 78.26%). In patients with definite TBM, the sensitivity of mNGS, AFB, PCR, and culture to detect MTB in the first CSF samples were 66.67, 33.33, 25, and 8.33%, respectively. The specificity of each method was 100%. Among the four negative mNGS cases (4/23, 17.39%), three turned out positive by repeated AFB stain. The agreement of mNGS with the total of conventional methods was 44.44% (8/18). Combination of mNGS and conventional methods increased the detection rate to 95.65%. One patient was diagnosed as complex of TBM and cryptococcal meningitis, in which AFB stain and cryptococcal antigen enzyme immunoassay were positive and the DNA of Cryptococcus neoformans was detected by mNGS.ConclusionOur study indicates that mNGS is an alternative method to detect the presence of mycobacterial DNA in CSF samples from patients with TBM and deserves to be applied as a front-line CSF test.

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Different next-generation sequencing pipelines based detection of tumor DNA in cerebrospinal fluid of lung adenocarcinoma cancer patients with leptomeningeal metastases.

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Different next-generation sequencing pipelines based detection of tumor DNA in cerebrospinal fluid of lung adenocarcinoma cancer patients with leptomeningeal metastases.

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