Abstract 4081: Analysis of low-grade serous ovarian cancer by long-read full length transcripts sequencing

Abstract Low grade serous ovarian carcinoma is a rare epithelial ovarian cancer that occurs more frequently in younger women. RNA sequencing (RNAseq) has been playing a pivotal role in understanding the molecular pathogenesis of the low-grade ovarian serous carcinoma (LGSOC). The quantification of gene expression with enough sequencing depth can be fairly accurate. However, the current short-read RNAseq approach is not very accurate in measuring individual transcript activity. This is because multiple transcripts from the same gene share high sequence similarity, which complicates the transcript mapping. As the accuracy of long-read sequencing technology is improving, long-read full transcript RNA sequencing is becoming an important tool for interrogating alternative transcripts and the discovery of gene fusion transcripts. In this study, we compare short-read RNAseq and long-read RNAseq from the same LGSOC patient samples. The RNA-Seq analysis for long-reads were performed by mapping sequencing reads to annotated reference human genome GrCh38 with minimap2. First, all annotated transcripts or genes were extracted including any annotated splice variants. Subsequently, the reads were mapped against all the transcripts as well as to the whole genome using minimap2. The distributing and counting the reads across genes and transcripts were calculated. We found that the expression level of transcripts in short-read in general is higher than that that of long-read. It is because short reads frequently could not be accurately mapped to the correct alternative transcripts from the same genes. When a read mapped to multiple transcripts, the short reads were randomly assigned to a transcript from the same gene. As a result, more than 100, 000 transcripts had at least one assigned read in the short-read RNAseq analysis, but less than 50, 000 transcripts had at least one assigned read in long-read RNAseq analysis. Long read with greater read-depth had better quantification accuracy for detecting specific transcripts. To identify potential gene fusions, the raw Oxford Nanopore long-read RNA-seq fastq files were corrected and polished with high sequencing quality short-read RNA-seq data by racon assembly tool. Those long-reads with unaligned ends of reads from a mapping generated by RNA-Seq analysis were re-mapped to the reference using the unaligned ends of reads. In this study, we identified several gene fusions including a gene fusion between NSUN4 and FAAH genes with at least seven long reads covering the breakpoint. FAAH encodes the enzyme fatty acid amide hydrolase and has been shown to be a metastasis suppressor in breast cancer previously. It is possible that the truncated FAAH protein because of gene fusion is involved in the metastasis of the LGSOC that we have analyzed. This could be another mechanism involved in the progression of LGSOC. Additional analysis of the long-read full transcript sequencing of LGSOC is in progress. Citation Format: Kwong-Kwok Wong, Yvonne Tsang, David M. Gershenson. Analysis of low-grade serous ovarian cancer by long-read full length transcripts sequencing [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 4081.