Introduction. Pancreatic cancer (PC) is emerging as one the most lethal types of cancer with only a 10% five-year survival rate [1]. This is attributed to the fact that most incidences of PC are diagnosed in advanced or metastatic stages [2]. In such cases, gemcitabine (GEM) remains one the first-line drugs used for treatment [2,3] but the efficacy of the treatment can fall short due to PC tumour heterogeneity and its acquired resistance to said chemotherapies [4]. A notable aspect of PC is the remarkable overexpression of aryl hydrocarbon receptor (AhR) mRNA and subsequently AhR itself with mRNA expression increasing on average 27-fold [5]. AhR is a ligand-dependent nuclear receptor which upon ligand binding activates the transcription of several metabolising enzymes which are implicated in tumour initiation and progression [6]. Due to poor future projections, low survival rates, limited treatment options and commonly observed resistance to drugs, it's imperative to discover ways to improve upon currently available treatments by means of modifying the expression of potentially involved genes.
Aim. The aim of this research was to investigate how AhR gene silencing can alter the effect of gemcitabine on cell metabolic activity and long-term colony formation of different PC cell lines.
Methods. PC cell lines BxPC-3 and Su.86.86 were transfected by lipofectamine mediated siAhR for 24 hours. Afterwards the cell lines were treated with IC50 doses of GEM (determined for each cell line separately) for 48 hours. After the treatment, short-term cytotoxic effect of GEM was determined by MTT assay and long-term effect was determined by seeding cells for clonogenic assay and subsequently growing them for 7 days.
Results. Treatment of the modified (siAhR) BxPC-3 cell line with the IC50 dose of GEM resulted in a statistically significant decrease in the short-term metabolic activity to 21,4 +/- 3,9 % in comparison to 50,6 +/- 10 % in unmodified control cells. As for the Su.86.86 cell line, AhR gene silencing had a statistically significant effect on metabolic activity in and of itself while also increasing cell sensitivity to GEM, however these results are inconclusive due to the mentioned responsiveness to silencing itself. In regards to long-term clonogenic assay, the IC50 dose of GEM decreased unmodified BxPC-3 colony formation to 18,3 +/-7,3% and Su.86.86 to 30,3 +/- 4,2% when compared to control cells. Silencing of AhR on its own resulted in a decrease of colony formation in the BxPC-3 line to 19,3 +/- 2,7 % and the Su.86.86 line to 27,7 +/- 4,9% when compared to control cells. AhR silencing in combination with GEM treatment also caused a decrease in colony formation capabilities compared to control cells: the BxPC-3 line dropping to 3,7 +/- 1,6 % and the Su.86.86 line to 11,3 +/- 3,2%. Furthermore, the Su.86.86 cell line was observed to be more resistant to GEM in both unmodified and transfected cells.
Conclusion. AhR expression modulation can possibly impact PC cell susceptibility to first-line drugs such as GEM which causes a detrimental effect on cell viability and long-term survival, however different responses of different cell lines show that the effect can vary depending on the profile of the cancer cells, therefore making reliable predictions difficult.