PO-496 Integrated omics- based approach to the identification of specific expression profiles associated with pten-loss

Introduction PTEN is a protein that restrains cells from growing unchecked; when it is absent or not functioning (as it often happens in prostate and colorectal cancer - CRC), cancer develops more easily and grows in a more aggressive and therapy-resistant fashion. However, just measuring PTEN mutational status or gene/protein levels in clinical samples may not be sufficient to assess its function, as its regulation is complex. Therefore, we analysed miRNA, transcriptomic, and proteomic profiles of isogenic CRC models differing for PTEN status, in order to identify a set of genes or proteins that are stably modulated by the presence or absence of a functional PTEN. Material and methods To identify biomarkers of PTEN loss of function and functional downstream effectors of PTEN activity, the isogenic CRC cell lines HCT116 (PTEN +/+ ) and HCT116 PTEN -/- (Horizon Discovery Ltd) were subjected to comprehensive miRNA profiling (Human miRNA Microarray Release 21, Agilent), transcriptomic analysis (NextSeq 500, Illumina), and semi-quantitative phosphoproteomic analysis (Antibody Microarray Service, KinexTM). Results and discussions miRNA profiling found 18 miRNAsdifferentially expressed in HCT116 PTEN +/+ and their HCT116 PTEN -/- counterpart: only miR-196a-5p is up-regulated in HCT116 PTEN -/- , whereas other 17 miRNAs were all down-regulated; several of these target miRNAs have been validated by qRT-PCR. RNA-Seq data were analysed via the RAP pipeline available at CINECA that relies on the Tuxedo suite (Tophat-Cufflinks-Cuffdiff) for read mapping, transcript abundance estimation and transcript or gene-based differential expression. RNA-Seq analysis showed 247 differentially expressed genes in HCT116 PTEN +/+ cells as compared to HCT116 PTEN -/- (159 up-regulated and 88 down-regulated in HCT116 PTEN -/- ) and clustering into 9 subgroups (including: tissue morphogenesis; negative regulation of phosphatase metabolic process; regulation of sequence-specific DNA binding transcription factor activity). qRT-PCR assay of 10 dysregulated genes was performed to validate the RNA-Seq data set. Proteomic analysis showed that 92 proteins (25 total proteins and 67 phospho-proteins) are differentially expressed in HCT116 PTEN +/+ and their HCT116 PTEN -/- counterpart, some of which have been validated by WB. Conclusion Our data identify differentially expressed sets of miRNA/genes/proteins between HCT116 PTEN +/+ and HCT116 PTEN -/- . Bioinformatics analysis to cluster these sets into functional PTEN-loss ‘signatures’ is ongoing.