Project description:Metformin is a drug used in the treatment of type 2 diabetes mellitus. Various studies have elucidated its anticancer properties. In this study, the effect of metformin on the differentiation and tumor microenvironment of colorectal cancer cells (CRC) was evaluated. For our study, we have used HCT116 colorectal cancer cell line and treated the cells with Metformin. Maximum tolerable non-toxic dose of metformin on HCT116 cells was determined by MTT assay. Cells were treated with 2.5 mM Metformin for 2 weeks. Analysis of apoptosis was done by flow cytometry using Annexin V / PI. CSC population was determined by flow cytometry using CSC markers CD44 and CD166. Metformin's ability to induce differentiation in CSC was assessed by analyzing Cytokeratin 20 (CK20) by flow cytometry and CDX1 (transcription factor for CK20), by RT-QPCR. Expression of Ki67 (proliferation marker) was done by RT-QPCR. RNA was isolated from 2.5 mM Metformin-treated and untreated cells populations. Microarray of untreated and 2.5 mM Metformin-treated RNA was done to study the whole genome transcriptomic changes.
Project description:In order to investigate the potential role of TP53 in regulating translation, HCT116 wild type and TP53 knockout cells were analyzed using both RNA sequencing, Ribosome sequencing and nascent proteome analysis. The cells were treated with 0.2 µg/ml Neocarcinostatin (NCS) to induce DNA damage and activate TP53.
Project description:Isoginkgetin (IGG) is a small molecule inhibitor of the spliceosome although the direct target remains elusive. Widespread failure to accurately remove introns poses a risk to cells and organisms through the potential production of aberrant mRNAs and proteins. The cellular responses to accumulation of these intermediates and/or the direct interference of spliceosome assembly itself may constitute a splicing stress but this is not well defined yet. We used oligonucleotide microarrays to assess genome wide changes in gene expression associated with exposure to IGG in HCT116 cells and an isogenic subline lacking the p53 tumor suppressor that responds to a variety of transcriptional stresses (Oncogene 18 (3), 583-592). Two of the 3 enriched pathways identified using PANTHER analysis of differentially expressed transcripts are linked to the ATF4 transcription factor and these effects are p53-independent.