Transcriptomic Analysis of Colon Cancer Cells Treated with an Anti-Diabetic Drug

Abstract

MASTER THESIS PROPOSAL for Jamila Hijazi Department of Biochemistry and Molecular Genetics

Transcriptomic analysis of colon cancer cells treated with an anti-diabetic drug

 Advisor: Dr. Georges Nemer  Co-Advisor: Dr. Pierre Khoueiry, Assistant Professor of Biochemistry & Molecular Genetics  Committee Members: 1. Dr. Nadine Darwiche, Professor of Biochemistry & Molecular Genetics 2. Dr. Assaad Eid, Associate Professor of Anatomy, Cell Biology & Physiological Sciences.

Abstract

Background: Metformin, The anti-diabetic drug which is widely used as a hypoglycemic agent in treating type-2 diabetes is also known for its anti-cancer effects. Data obtained from in-vitro and in-vivo studies showed that metformin influences cancer progression since it affects cell cycle arrest and apoptosis. Most studies refer this anti-proliferation mechanism to the inhibition of mTor by metformin; however the genetic pathway analysis underlying the downstream targets is still rarely reported.

Aims: In this study, we aim to unravel the genetic pathways using in-vitro cell model, and to examine how metformin affect cell cycle and proliferation in colon cancer cells with respect to p21 and p53 status.

Methods: Four different metformin concentrations were used (1mM, 2mM, 5mM and 10 mM) on the HCT116 colon cancer cell line and its genetic modified forms HCT-116 p53 null and p21 null. To assess Percentage of cell viability MTT assay was applied at 24hr and 48hr of treatment. Cell cycle and apoptosis rate was measured using the flow cytometer propidium iodide based assay.

Finally, RNA-seq was performed on HCT116 after 24hr of metformin treatment and the results were analyzed using Galaxy. Pathway analysis was undertaken using the Pathway studio software, and significantly differentially expressed genes were confirmed by qPCR.

Preliminary results showed a significant decrease in the percentage of cell viability between the control and the treated cancer cells in a dose and time response manner. Analysis of cell cycle has shown an increased in the subG0 phase (apoptosis phase)especialy in the HCT116 p21-/-. Transcriptomic analysis revealed more than 3,000 significantly down-regulated genes and 1,200 up-regulated genes (p<0.05).

Conclusion: metformin shows high potential in inhibiting colorectal cancer proliferation and in inducing apoptosis through increasing cell death at the level of SubG0 especially in the absence of p21 gene as p53 could be an important player in directing cells towards apoptosis but it’s not alone. Metformin affected many molecular pathways implicated in colorectal cancer but still more of NGS should be done at different levels.