DNMT1: An emerging target in the treatment of invasive urinary bladder cancer

Objectives: More than 14,000 people die from invasive urothelial carcinoma (iUC) of the urinary bladder each year in the USA, and more effective therapies are needed. Naturally occurring canine iUC very closely resembles the disease in humans and serves as a highly relevant translational model for novel therapy of human iUC. Work was undertaken to identify new targets for anticancer therapy in dogs with the goal of translating successful therapeutic strategies into humans with iUC. Materials and methods: Microarray expression analyses were conducted on mRNA extracted from canine normal bladder (n = 4) and iUC tissues (n = 4) using Genome Array 1.0 and analyzed by GeneSpring GX 11, with the stringency of P < 0.02 and a >= 2-fold change. The genes thus identified were further analyzed for functional and pathway analysis using Protein ANalysis THrough Evolutionary Relationships (PANTHER) Classification System. In selecting genes for further study, consideration was given for evidence of a role of the gene in human iUC. From these analyses, DNA methyltransferase 1 (DNMT1) was selected for further study. Immunohistochemistry (IBC) of canine normal bladder and iUC tissues was performed to confirm the microarray expression analyses. The effects of targeting DNMT1 in vitro was assessed through MTT assay and Western blot of canine iUC cells treated with 5-azacitidine (5-azaC) and trichostatin A (TSA). Results: DNMT1 was expressed in 0 of 6 normal canine bladder samples and in 10 of 22 (45%) canine iUC samples. The proliferation of canine iUC cells was inhibited by 5-azaC (at concentrations >= 5 mu m) and by TSA (at concentrations >= 0.1 mu m). Western blot results were supportive of DNMT1-related effects having a role in the antiproliferative activity. Conclusions: Microarray expression analyses on canine tissues identified DNMT1 as a potentially "targetable" gene. Expression of DNMT1 in canine iUC was confirmed by IHC, and in vitro studies confirmed that drugs that inhibit DNMT1 have antiproliferative effects. These findings are similar to those recently reported in human iUC and are also in line with results of a preclinical (prehuman) trial of 5-azaC in dogs with naturally occurring iUC. DNMT1 has excellent potential as a target for iUC therapy in humans. (C) 2013 Elsevier Inc. All rights reserved.