Inden J. Pharmacol Rev 2001; 53: 5272. two. Poulsen SA, Quinn RJ. Bioorg Med Chem 1998; 6: 6191. three. Colotta V, Catarzi D, Varano F, Cecchi L, Filacchioni G et al. J Med Chem 2000; 43: 31184.SpringerCommunications2-Chloro-20 -Deoxyadenosine-Induced Apoptosis of Human Cancer Cells Bearing a Mutated p53 Isoform: Function of MAP KinasesStefania Ceruti, Alessia Mazzola and Maria P. AbbracchioLaboratory of Molecular and Cellular Pharmacology of Purinergic Transmission – Department of Pharmacological Sciences, University of Milan – by way of Balzaretti 9 – 20133 – Milan, Italy [email protected] The biochemical pathways controlling cell death are usually mutated and/or defective in cancer cells, contributing to the improvement of resistance to chemotherapy. As a result, for any given anti-cancer agent, it’s crucial to confirm irrespective of whether it might efficiently induce cell death in cancerous cells carrying mutations of different apoptotic pathways. We have previously demonstrated that human astrocytoma ADF cells are resistant to agents activating the intrinsic pathway of apoptosis due the presence of a mutated caspase-9 isoform (1). Nevertheless, the anti-cancer agent 2-chloro-20 -deoxyadenosine (Cladribine, 2CdA) induces cell cycle block and continues to be capable to properly kill these cells by 49843-98-3 In Vivo recruiting an atypical pathway of cell death, with caspase-2 acting as an “initiator” caspase, followed by caspase-3 (2). To better characterize 2CdA-activated apoptotic pathway in ADF cells, in this study we have focussed our attention around the tumor suppressor protein p53 which can be mutated in more than 50 of human cancers. By cloning and sequencing the p53 isoform expressed by ADF cells, we detected a single nucleotide mutation in position 797 which translated into a G-to-E substitution at position 266 which belongs towards the DNA-binding domain of your p53 protein. Considering the fact that phosphorylation from the Ser15 residue of the p53 protein represents one of by far the most frequent post-translational modification controlling its proapoptotic activity, we’ve analyzed the value of this phosphorylation reaction in cells exposed for a variety of time periods to 2CdA by implies of a specific antibody directed against phosphorylated p53. An incredibly fast apperance from the protein band 88899-55-2 Biological Activity corresponding to phospho-Ser15 p53 was observed, starting from a 30 min and declining following 3 hours of incubation with the adenosine analog. P53 phosphorylation was completely prevented by 2-deoxy-cytidine, which competes with 2CdA for its intracellular phosphorylation for the corresponding chloro-deoxy nucleotides. Certainly, co-incubation with SP600125, a selective inhibitor of c-jun-terminal kinases 1 and 2 (JNK1/2), entirely blocked p53 phosphorylation, whereas inhibitors of other members with the MAP kinase family members of enzymes (e.g., ERK1/2 and p38) had no impact. SP600125 was also in a position to drastically reduce the percentage of apoptosis induced by 2CdA. Nonetheless, p53 phosphorylation and 2CdA-induced apoptosis are not correlated to each other, considering the fact that !-pifithrin, which selectively inhibits p53 transcriptional activity downstream of post-translational modifications, was totally ineffective in preventing cell death. Taken collectively, these outcomes suggests that a JNK-dependent but p53-independent pathway of death is recruited by 2CdA. JNK1/2 had been also involved inside the 2CdA-induced cell cycle block at G0/G1 phases and 370-86-5 Epigenetics enhanced expression of genes controlling cell cycle progression (e.g., p21, GADD45A) as demonstrated by microarray evaluation.