Metallothionein isoform 3 as a potential biomarker for human bladder cancer

The goal of the present study was to determine if the expression of metallo thionein isoform 3 (MT-3) might serve as a biomarker for human bladder canc er. To accomplish this goal, we defined the localization and expression of MT-3 protein and mRNA using fresh and archival biopsy specimens obtained fr om patients undergoing differential diagnosis for a variety of bladder diso rders. We used immunohistochemistry, immunoblot, and RT-PCR analysis to def ine the localization and expression of MT-3 protein and mRNA. Immunohistoch emical analysis disclosed no immunoreactivity for MT-3 in normal bladder ce lls. The absence of MT-3 expression in the normal bladder was further confi rmed by demonstrating that MT-3 mRNA could not be detected using reverse tr anscriptase-polymerase chain reaction (RT-PCR) or MT-3 protein using immuno blot. Immunohistochemistry also disclosed no immunoreactivity for MT-3 in a rchival biopsy specimens from patients with interstitial cystitis and relat ed disorders. Immunohistochemical analysis demonstrated that MT-3 was expre ssed in carcinoma in situ (CIS), high-grade bladder cancer, low-grade bladd er cancer, and dysplastic lesions. MT-3 immunostaining was intense in both CIS and high-grade bladder cancer, and low to moderate in low-grade bladder cancer and dysplastic lesions. We determined MT-3 mRNA expression in a sub set of these bladder cancer specimens; expression was elevated as compared to that of the housekeeping gene, beta-actin. The cDNA from the RT-PCR reac tion primed for MT-3 contained a FokI restriction site, a site unique for M T-3 as compared to other MT family members. In conclusion, this study demon strates that MT-3 is up-regulated in human bladder cancer and that this up- regulation increases with increasing tumor grade. The finding that MT-3 exp ression is minimal in normal bladder suggests that MT-3 might be developed into an effective biomarker for bladder cancer.