Systematic Identification Of Long Noncoding RNAS Of Platinum Sensitivity And Resistant In Ovarian Cancer Based On RNA-Seq And Its Differential Expression Analysis And Study

Long non-coding RNAs, lncRNAs, which exists in the nucleus or cytoplasm with a transcript length between 200nt and100kb, does not or rarely code for protein itself. Evidence is accumulating that lncRNAs and cancer are closely related, and there are differentially expressed in normal and tumor cells. The abnormal expression of lncRNAs may play an important role in the occurrence of tumor. Some lncRNAs can promote carcinogenesis, which is highly expressed in the ovarian cancer; some can inhibit the tumor. Ovarian cancer is a kind of malignant tumor, which grow rapidly and spread easily. Through operation combined with MDT based on platinum,60% to 80% of patients by first-line treatment can relieve completely. The sensitivity of platinum is an important reference for the salvage treatment, which demonstrates a growing role in the clinical application. This study based on RNA-Seq data by bioinformatic approaches systematically identify lncRNAs of platinum sensitivity and resistant in ovarian cancer and analyze their differential expressions and related functions. The main contents are as following1) 12 sample data with RNA-Seq paired-end and platinum resistant and sensitive conditions in ovarian cancer were downloaded from ArrayExpress based on Immila platform. By FastQC, FASTX-Toolkit and Kmer histogram evaluated its quality control; according to the characteristics of these data, a reasonable method of data preprocessing was developed.2) Based on the preprocessing of data, transcriptome reconstruction of the read data was conducted, including mapping and assembly:The tophat2 were first used for reads mapping to the human reference genomics, the ratio ofmappinged reads was between 93.6% and 94.4%, the ratio of uniquemapping on paired-end reads was 79.20%～83.6%. Then using thecufflinks to assemble the mapping read, and 124,361 transcripts were obtained which distributed in 78,900 different Locis.3) A method about systematic identification of long noncoding RNAs based on RNA-Seq in ovarian cancer with platinum sensitivity and resistant was presented. The coding transcriptswere filtered according to transcript length, exon number and coverage of maximum read respectively; and filtered the protein coding transcripts by using UCSC、RefSeq、Ensembland Encode4 database annotation; then using coding potential tools predicted the potential for the rest of transcript encoding, which became the candidate lncRNAs transcription; Finally the lncRNAs were estimated using HMMER-3 for the protein domains, to get the potential lncRNAs transcription, and 1325 of lncRNAs in ovarian cancer with platinum sensitive and resistant were identified, including 1162 known and 163 novel lncRNAs.4) The lncRNAs of sensitivity and resistant in ovarian cancer were identified by analyzing the expression difference. We found that there were 46 lncRNAs showing significant differential expression, including 6 novellncRNAs and 40 known ones. To further study the function of both sensitivity and resistant of lncRNAs in ovarian cancer, we also analysed these 46 significant expressed lncRNAs by GO enrichment and pathway analysis. Results showed that differential expressed lnRNAs associated with bingding(organic cyclic compound binding、ion binding), intra uterine growth and immune process. Based on the analysis of lncRNAs pathway, the result showed that the interference of pathway includes nucleotide metabolism, calcium signaling pathway, pertussis disease, systemic lupus erythematosus, biological synthesis of secondary metabolism and cancer related transcription disorders.5) Quantitative RT-PCR was performed on the expression level for the differentially expressed lncRNAs. We selected three lncRNAs (two from novel lncRNAs, one from known lncRNAs), and verified its expression value,the result showed expression fitted well with analysis results.Based on the above analysis, a systematic method to identify lncRNAs in ovarian cancer with platinum sensitive and resistant was established, by the analysis of significantly expressed lncRNAs, we found lncRNAs had relationship with bingding, intra uterine growth, immune process and cancer related transcriptional dysregulation. In the future, if we could vertify the significant differential lncRNAs through different experimentsorganization, it may be used as a biomarker in diagnosis of resistance of ovarian cancer.