Transcriptional Profile of HCV Replicon Cells after Treatment with Acetylsalicylic Acid.

OBJECTIVE: It has been demonstrated in vitro that acetylsalicylic acid (ASA) treatment halves hepatitis C virus (HCV) expression in hepatocarcinoma cells. However, the signaling pathway that promotes this ASA-induced antiviral effect has not yet been identified. AIM: The aim of this work was to identify alterations in the transcriptional profile of Huh-7-HCV-subgenomic replicon cells with vs. without ASA treatment. This comparison sheds light onto the signaling pathways and molecular mechanisms involved in the antiviral effects of ASA. METHODS: Human hepatocellular carcinoma (Huh-7) cells that express non-structural HCV proteins (Huh-7-HCV-replicon cells) were exposed to 4 mM ASA for 0, 24, 48, and 72 hours. Total RNA was isolated, and cDNA was synthesized. Transcripts were then tagged with biotin and purified. Thereafter, they were fragmented and hybridized on HG-U133 Plus 2 Gene Expression chips. Hybridization signals were captured using a GeneChip 3000 7G Scanner and analyzed via Expression Console and dChip Software. RESULTS: When exposed to ASA, hepatocarcinoma cells with non-structural HCV proteins were found to differentially regulate genes with oxidative roles in the cell. The most upregulated genes were interleukin 8 (IL-8), cytochrome P450 (CYP450), and metallothioneins (MTs), while the most downregulated genes were ribonucleotide reductases (RRs). CONCLUSION: These results show that ASA modulates the expression of genes with antioxidant functions. This suggests that ASA induces a remodeling of the antioxidant microenvironment, which may in turn interfere with the replication of HCV.

Down regulation of tumour biomarkers in colon cancer cells with IRNA PFK-1 plus metformin.

PURPOSE: Metformin has been widely used for the treatment of Type 2 Diabetes Mellitus (T2DM), hyperglycemia and polycystic ovarian syndrome. Recent studies have suggested the potential of this substance as a cancer chemopreventive agent. We evaluated the antitumoral effect of iRNA-PFK-1 and the combined therapy iRNA-PFK-1 + metformin in RKO p53-positive cells. METHODS: mRNA levels of tumor suppressor genes AMPK, APC, and c-MYC, KRAS oncogenes were measured by qRT-PCR in RKO cells treated with 25 µM metformin alone or combined with iRNA-PFK-1, to evaluate the effect of both treatments. RESULTS: At 72 h after treatment with either 25 µM metformin, 150 nM iRNA-PFK-1, or the combined treatment, the transcriptional levels of these biomarkers were decreased by ~73% (p˂0.05), ~99.9%, (p˂0.01), and ~76% (p˂0.05), respectively. CONCLUSION: These in vitro results support the potential therapeutic role of metformin and PFK-1 in the treatment of colon cancer via down-modulation of the expression of several important cancer biomarkers.