Abstract PD4-2: Whole exome and transcriptome sequencing of 120 primary breast cancer to discover novel therapeutic target

Abstract

Abstract Introduction: Many somatic mutations, structural alterations, and gene expression changes are causally implicated in oncogenesis and tumor progression, and as a result, affect clinical outcome. Although majority of breast cancer patients have benefits from therapeutics targeting tumor biology, such as estrogen receptor and HER-2, still many patients suffer from disease recurrence and metastasis. More kind of specific target therapies are needed, especially for hormone-resistant tumor and triple-negative breast cancer. Materials and Method: To find novel therapeutic target in breast cancer, here we examine the both whole exome and whole transcriptome of fresh-frozen primary breast cancer tissues from 120 patients whose clinical, pathological, and survival data are available. Patients with Stage IV disease or who received neoadjuvant chemotherapy were excluded. 36 patients had distant metastasis within 5 years from surgery, and 84 patients were NED at least 5 years. RNA and DNA were extracted and qualities were assessed in all samples. Exome and transcriptome sequencing were done using NGS technology (Illumina HiSeq 2000). As a control, exome sequencing was done for 93 normal DNA from matched patients. Single nucleotide variations (SNV) identified in cancer samples on exonic region, nonsynonymous SNV or stop gain/loss, whose quality ≥20, and not found in 93 normal samples were included. SNVs registered in dbSNP135_common or 1000 genome allele frequency >0.001 were excluded.Results and Discussion: We identified 11,684 putative somatic mutations in 7,373 genes. Of them, 6,547 were deleterious or damaging mutation by Provean or SIFT analysis. Mutations were found in potential drug target genes, such as PIK3CA(25), PTEN(3), AKT1(3), ALK(3), ROS1(2), FGFR4(3), FGFR3(2), ERBB2(2), and IDH1(1) etc. In a pathway analysis, mutations in insulin signaling pathway were most dominant. We hypothesized that driver gene and therapeutic target has to have recurrent mutation and gene expression at least more than average expression. We calculated expression “Volume” according to the median normalized FPKM value of individual gene's RNA-seq data. With a cut-off of 3 or more mutations in each gene, 1,116 genes were selected. After the filtering of Volume<0.3, 696 genes were selected. Finally, 55 genes were selected which are druggable or potentially druggable using drug database (DrugBank, TOCRIS, Ingenuity) and Pubmed. DriverNet analysis result was also considered for the selection. All 342 tumor suppressor genes were filtered out. Interestingly, 18 of the 55 were genes involved in metabolism (fatty acid, glucose, amino acids). 12 were kinases and 4 were involved in insulin pathway. Excluding the previously confirmed therapeutic target, PIK3CA, AKT1, and NOTCHs, and considering the patients’ clinical data, our primary candidates for hormone-resistant breast cancer were NQO2, CELSR1, GLUD2, MYH9, PSMD2, NADK, IRS2, MAP3K5, and for triple-negative breast cancer were HSPG2, PHGDH, MYLK, etc. Validation with Sanger sequencing and functional study is on-going. Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr PD4-2.