Xuorouracil on Silybin B apoptosis was examined in vitro using PARP cleavage and TUNEL. Finally, the eVectiveness of combining PXD101 and 5 Xuorouracil in vivo was tested using both HT 29 and HCT116 xenograft models. Results Synergistic inhibition of proliferation and clonogenicity was obtained when HCT116 cells were incubated with PXD101 and 5 Xuorouracil. 5 Xuorouracil combined with PXD101 also increased DNA fragmentation and PARP cleavage in HCT116 cells. Incubation with PXD101 down regulated thymidylate synthase expression in HCT116 cells. In vivo studies, using mouse HT29 and HCT116 xenograft models, showed improved reductions in tumour volume compared to single compound, when PXD101 and 5 Xuorouracil were combined.
Conclusions PXD101 and 5 Xuorouracil synergistically combine in their anti tumour eVects against colon cancer cells in vitro and show enhanced activity when combined in vivo. Based on the results presented herein, a rationale for the use of PXD101 and 5 Xuorouracil in combination in the clinic has been demonstrated.In PARP Inhibition vitro studies have established that in 5 FU resistant cell lines, the levels Fesoterodine price of TS rise dramatically and this is a major contributing factor towards the mechanism of resistance . Elevated levels of tumour TS have also been repeatedly demonstrated to be a major contributor towards resistance to 5 FU, with high levels of TS being predictive of a poor response . Gene expression is controlled in part by a class of enzymes known as histone deacetylases . HDACs are zinc dependent hydrolases that control remodelling of chromatin by deactylation of speciWc lysine residues on histone tails .
The deacetylase action of HDACs has the eVect of condensing chromatin and therefore restricting access to the DNA for nuclear proteins such as Erlotinib ic50 transcription factors, leading to alterations in gene expression. Histone deacetylase inhibitors are currently being developed as anti tumour agents and have been shown to inhibit the growth and induce apoptosis of hyper proliferating cancer cells. Expression proWling of cells treated with HDACi in vitro showed that under these conditions genes are both up and down regulated, at least at the mRNA level . Up to 5% of the total genome has been shown to be regulated by HDACi using microarrays, although the exact Wgure appears to be dependent on the HDACi and cell line used in the study .
Altered genes included those required for cellular proliferation, signs signal transduction, metabolism and metastasis, as well as others. This data has encouraged the investigation of combining HDACi and compounds that have their molecular target regulated by HDACi. Indeed, it has been demonstrated that combinations of HDACi with well established chemotherapeutics can synergise with their antitumour eVects . We have examined in detail the eVects of PXD101 in combination with 5 FU on colorectal tumour cell proliferation and apoptosis, both in vitro and in vivo. PXD101 is an HDACi that shows nM and sub M potency in HDAC biochemical and anti proliferative in vitro assays respectively . PXD101 is currently undergoing phase I/II clinical evaluation for the treatment of multiple cancers types including T cell lymphoma, multiple myeloma, ovarian and colorectal. Data provided here establishes a rationale for the use of HDACi.