Abstract
BackgroundIt has been reported that the histone deacetylase inhibitor (iHDAc) trichostatin A (TSA) induces an increase in MDR1 gene transcription (ABCB1). This result would compromise the use of iHDACs in combination with other cytotoxic agents that are substrates of P-glycoprotein (Pgp). It has also been reported the use of alternative promoters by the ABCB1 gene and the existence of a translational control of Pgp protein. Finally, the ABCB1 gene is located in a genetic locus with the nested gene RUNDC3B in the complementary DNA strand, raising the possibility that RUNDC3B expression could interfere with ABCB1 alternative promoter regulation.MethodsA combination of RT-PCR, real time RT-PCR, Western blot and drug accumulation assays by flow cytometry has been used in this study.ResultsThe iHDACs-induced increase in MDR1 mRNA levels is not followed by a subsequent increase in Pgp protein levels or activity in several pancreatic and colon carcinoma cell lines, suggesting a translational control of Pgp in these cell lines. In addition, the MDR1 mRNA produced in these cell lines is shorter in its 5′ end that the Pgp mRNA produced in cell lines expressing Pgp protein. The different size of the Pgp mRNA is due to the use of alternative promoters. We also demonstrate that these promoters are differentially regulated by TSA. The translational blockade of Pgp mRNA in the pancreatic carcinoma cell lines could be related to alterations in the 5′ end of the MDR1 mRNA in the Pgp protein expressing cell lines. In addition, we demonstrate that the ABCB1 nested gene RUNDC3B expression although upregulated by TSA is independent of the ABCB1 alternative promoter used.ConclusionsThe results show that the increase in MDR1 mRNA expression after iHDACs treatment is clinically irrelevant since this mRNA does not render an active Pgp protein, at least in colon and pancreatic cancer cell lines. Furthermore, we demonstrate that TSA in fact, regulates differentially both ABCB1 promoters, downregulating the upstream promoter that is responsible for active P-glycoprotein expression. These results suggest that iHDACs such as TSA may in fact potentiate the effects of antitumour drugs that are substrates of Pgp. Finally, we also demonstrate that TSA upregulates RUNDC3B mRNA independently of the ABCB1 promoter in use.
Highlights
It has been reported that the histone deacetylase inhibitor trichostatin A (TSA) induces an increase in MDR1 gene transcription (ABCB1)
Effect of histone deacetylase inhibitors on Pgp mRNA expression in different pancreatic carcinoma cell lines To determine iHDACs effects in MDR1 mRNA expression in human pancreatic carcinoma cell lines, we analyzed the level of MDR1 mRNA by real time RT-PCR in IMIM-PC-1, IMIM-PC-2 and RWP-1 human pancreatic carcinoma cell lines in the presence and absence of TSA and Suberoylanilide Hydroxamic Acid (SAHA)
Despite this increase in MDR1 mRNA levels, we have previously reported that TSA and other iHDACs are able to inhibit cell growth and to induce apoptosis in these pancreatic carcinoma cell lines [24]
Summary
It has been reported that the histone deacetylase inhibitor (iHDAc) trichostatin A (TSA) induces an increase in MDR1 gene transcription (ABCB1) This result would compromise the use of iHDACs in combination with other cytotoxic agents that are substrates of P-glycoprotein (Pgp). Several mechanisms could be involved in the acquisition of this phenotype, the role of two different membrane proteins, P-glycoprotein (Pgp) and multidrug resistance associated protein (MRP), has been well established [2,3,4]. Both proteins are members of the same ATP-binding cassette (ABC) superfamily of transport proteins. It is clear that Pgp has several functions in different cells and tissues
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